JP7216157B2 - Methods for treating or preventing asthma by administering an IL-4R antagonist - Google Patents

Description

RELATED APPLICATIONS This application is based on U.S. Provisional Application No. 61/691,625 (filed Aug. 21, 2012); U.S. Provisional Application No. 61/758,097, filed Jan. 29, 2013; /761,279, filed February 6, 2013; U.S. Provisional Application No. 61/783,796, filed March 14, 2013; U.S. Provisional Application No. 61/805,797, filed March 27, 2013 and claims the benefit of priority from French Application No. 1356994, filed July 16, 2013. The contents of each of the aforementioned applications are hereby incorporated by reference in their entirety.

FIELD OF THE INVENTION This invention relates to the treatment and/or prevention of asthma and related conditions. More particularly, the invention relates to administration of interleukin-4 receptor (IL-4R) antagonists to treat or prevent asthma in patients in need thereof.

BACKGROUND Asthma is a chronic inflammatory disease of the airways characterized by airway hyperreactivity, acute and chronic bronchoconstriction, airway edema, and mucus plugging. The inflammatory component of asthma is thought to involve many cell types, including mast cells, eosinophils, T lymphocytes, neutrophils, and epithelial cells and their biological products. Asthmatics most frequently exhibit wheezing, shortness of breath, coughing, and chest tightness. For most patients, a regimen of controller therapy and bronchodilator therapy provides adequate long-term management. Inhaled corticosteroids (ICS) are considered the "gold standard" in the management of asthma symptoms, and inhaled beta2-agonists are the most effective bronchodilators currently available. Studies have shown that combination therapy with ICS and an inhaled long-acting beta2-agonist (LABA) provides better asthma control than high-dose ICS alone. As a result, combination therapy has become the treatment of choice for subjects not managed with low doses of ICS alone.

Nevertheless, it is estimated that 5% to 10% of the population with asthma has symptomatic disease despite maximum recommended treatment with a combination of anti-inflammatory and bronchodilator drugs. In addition, this severe asthma population accounts for up to 50% of total medical costs due to hospitalizations, use of emergency services, and emergency visits. As many of these patients are poorly responsive to ICS due to multiple cellular and molecular mechanisms, there is an unmet need for new treatments in this severe asthma population. In addition, long-term adverse effects of systemic and inhaled corticosteroids on bone metabolism, adrenal function, and growth in children have led to attempts at minimal corticosteroid utilization. Although the majority of asthma patients are reasonably well controlled with current treatments, patients with severe corticosteroid-resistant asthma have few therapeutic treatment options that can adequately manage their disease. The result of non-response to treatment or lack of adherence to treatment is loss of asthma control and ultimately asthma exacerbation.

One reason for the poor response to pharmacotherapy in some patients with severe asthma may be the heterogeneity of the disease. There is growing interest in understanding these unique phenotypes, as targeted therapies are more likely to be successful in patients with similar underlying pathologic features. Recent therapeutic approaches in asthma have focused on trying to manage the T helper cell-2 response. Upregulation of interleukin-4 (IL-4) and interleukin-13 (IL-13) has been shown to be an important inflammatory component of asthma disease progression.

Accordingly, there is a need in the art for new targeted therapies for the treatment and/or prevention of asthma.

SUMMARY OF THE INVENTION According to one aspect of the invention, methods are provided for reducing the incidence of asthma exacerbations in a subject in need thereof. In a related aspect, methods are provided for improving one or more asthma-related parameters in a subject in need thereof. In yet another aspect of the invention, methods are provided for treating asthma, eg, moderate to severe eosinophilic asthma, in a subject in need thereof.

A method featured in the invention comprises administering to a subject a therapeutically effective amount of a pharmaceutical composition comprising an interleukin-4 receptor (IL-4R) antagonist. According to a particular embodiment, the IL-4R antagonist is an antibody or antigen-binding fragment thereof that specifically binds IL-4R. Exemplary anti-IL-4R antibodies that can be used in the context of the methods of the invention are described elsewhere herein, including in Example 1. For example, in one embodiment, the IL-4R antagonist is an antibody or antigen-binding fragment thereof that specifically binds IL-4R and has the heavy chain variable region (HCVR) and light chain variable region of SEQ ID NOs: 162 and 164, respectively. Includes heavy and light chain (complementarity determining region) CDR sequences from the region (LCVR).

In one embodiment, for reducing the incidence of one or more asthma exacerbations in a subject in need thereof by administering an antibody or antigen-binding fragment thereof that specifically binds IL-4R. A method is provided. An asthma exacerbation can be one or more of the following: (a) 30% or more reduction from baseline in morning peak expiratory flow (PEF) for 2 consecutive days; (b) 2 consecutive days. , 6 or more additional seizure reliever puffs of albuterol or levalbuterol in 24 hours (compared to baseline); and (c)(i) systemic (oral and/or parenteral) steroids treatment, or (ii) an increase in inhaled corticosteroid to at least four times the last dose received prior to discontinuation, or asthma exacerbation requiring hospitalization.

In various embodiments, the method for ameliorating one or more asthma-related parameters comprises administering to a subject in need thereof a therapeutically effective amount of an IL-4R antagonist, wherein asthma-related Parameter improvement is defined as one of the following: increase from baseline in FEV1; increase from baseline in AM PEF; increase from baseline in PM PEF; a decrease from baseline; a decrease from baseline in nocturnal alertness; and/or a decrease from baseline in SNOT-22 score. Examples of asthma-related parameters include: (a) forced expiratory volume in 1 second (FEV1); (b) peak expiratory volume including morning PEF (AM PEF) and evening PEF (PM PEF). (c) use of inhaled bronchodilators such as albuterol or levalbuterol; (d) five-item Asthma Control Questionnaire (ACQ5) score; (d) nocturnal awakening; and (e) 22-item Sino-Nasal Outcome Test (SNOT-22) score. In one embodiment, the improvement in the asthma-related parameter is an increase from baseline in FEV1 of at least 0.10 L. In one embodiment, the improvement in the asthma-related parameter is an increase from baseline in AM PEF of at least 10.0 L/min. In one embodiment, the improvement in the asthma-related parameter is an increase from baseline in PM PEF of at least 1.0 L/min. In one embodiment, the improvement in the asthma-related parameter is a reduction from baseline of at least 1 puff per day of albuterol/lev-albuterol use. In one embodiment, improvement in an asthma-related parameter is a reduction from baseline of at least 0.5 points in ACQ5 score. In one embodiment, an improvement in an asthma-related parameter is a reduction from baseline in nocturnal awakenings of at least 0.2 times per night. In one embodiment, improvement in an asthma-related parameter is a reduction in SNOT-22 score of at least 5 points from baseline.

The invention also provides a method for reducing the incidence of asthma exacerbations or improving one or more asthma-related parameters in a subject in need thereof, wherein the method comprises: A single initial dose of a pharmaceutical composition comprising an IL-4R antagonist (eg, an anti-IL-4R antibody or antigen-binding fragment thereof), followed by one or more secondary doses of a pharmaceutical composition comprising an IL-4R antagonist to a subject in need thereof sequentially. A pharmaceutical composition comprising an IL-4R antagonist can be administered subcutaneously, intranasally, or intravenously to a subject in need thereof.

According to certain embodiments, the present invention provides methods for reducing the incidence of asthma exacerbations or improving one or more asthma-related parameters in a subject in need thereof. wherein the method comprises administering to the subject from about 75 to about 300 mg of a pharmaceutical composition comprising an antibody or antigen-binding fragment thereof that specifically binds IL-4R. According to this aspect, the pharmaceutical composition can be administered to the subject at a dosing frequency of, for example, once a week.

The invention further provides for selecting a subject exhibiting one or more symptoms or signs of asthma and providing a pharmaceutical composition comprising an IL-4R antagonist (eg, an anti-IL-4R antibody or antigen-binding fragment thereof). It includes methods of treating asthma (e.g., eosinophilic asthma, moderate-to-severe eosinophilic asthma, etc.) by administering to a patient, wherein the subject has the following symptoms or signs of asthma: (1) subject is on fluticasone/salmeterol combination therapy (250/50 μg twice daily (BID) or 500/50 μg BID) or budesonide/formoterol combination therapy (160/9 μg BID or (2) the subject has blood eosinophils greater than or equal to 300 cells/μL; (3) the subject has sputum eosinophils greater than or equal to 3%; (4) the subject has elevated levels of IgE, thymus and activation regulation chemokines; (TARC), eotaxin-3, carcinoembryonic antigen (CEA), YKL-40, or periostin; (5) the subject has elevated levels of fractional exhaled nitric oxide (FeNO); and/or (6) the subject has a 5-item Asthma Control Questionnaire (ACQ5) score greater than or equal to 1.0.

Embodiments featured in this invention relate to the above treatment methods, further comprising administration of a second therapeutic agent in combination with the IL-4R antagonist. The second therapeutic agent can be administered to a subject in need thereof before, after, or concurrently with the IL-4R antagonist. Exemplary second therapeutic agents include, but are not limited to, one or more of the following in combination: IL-1 inhibitors, IL-5 inhibitors, IL-8 inhibitors, IgE inhibitors, Tumor necrosis factor (TNF) inhibitors, corticosteroids, long-acting beta2-agonists, and leukotriene inhibitors.

In another aspect, the invention provides a method of reducing or eliminating an asthma patient's dependence on background asthma therapy, wherein the method is uncontrolled or partially controlled with background asthma therapy. selecting patients with moderate to severe asthma; administering a defined dose of an IL-4R antagonist to the patient while maintaining the patient's background asthma therapy; This includes gradually decreasing the dosage of one or more components of the background therapy over the treatment period. In certain embodiments, background therapy comprises inhaled corticosteroids (ICS), long-acting beta-agonists (LABA), or a combination of ICS and LABA. In some embodiments, background therapy is tapered or discontinued over a period of 2-8 weeks. In some embodiments, one component of background therapy is eliminated after the initial treatment phase. In one embodiment, the background therapy is gradually decreased over the subsequent treatment period.

In yet another aspect, the invention has elevated levels of biomarkers such as thymic and activation-regulated chemokines (TARC), IgE, eotaxin-3, periostin, carcinoembryonic antigen (CEA), or YKL-40. or by selecting patients with increased levels of fractional exhaled nitric oxide (FeNO); and administering to the patient a therapeutically effective amount of an IL-4R antagonist; and provides a method for treating moderate to severe asthma.

In another aspect, the invention provides, for example, (a) the level of expression of one or both of TARC or eotaxin-3, or IgE in a biological sample obtained from a subject prior to treatment with an IL-4R antagonist. (b) determining the expression level of the biomarker in a biological sample obtained from the subject after treatment with the IL-4R antagonist; (c) determining in step (a) comparing the determined expression level to the level in step (b), and (d) if the level determined in step (b) is lower than the level determined in step (a), the treatment is effective. moderate to moderate in the subject by concluding or concluding that treatment is not effective if the level determined in step (b) is the same or higher than the level determined in step (a) A method for monitoring the efficacy of treatment for severe asthma is featured.

In one embodiment, the biomarker is FeNO, and treatment with an IL-4R antagonist is determined to be efficacious if FeNO levels decrease following administration of the antagonist.

The expression level of the biomarker can be determined, for example, after 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks or more after administration of the IL-4R antagonist, and the expression level prior to administration of the antagonist. can be compared with The dose or dosing regimen of the IL-4R antagonist (eg, anti-IL4R antibody) can be adjusted after that determination. For example, if biomarker expression does not decrease within 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, or more after administration of the antagonist, treatment with the antagonist can be discontinued; Or the dose of the antagonist can be increased. If biomarker expression decreases following administration of an antagonist, the dosage of that antagonist can be maintained or decreased, eg, to identify a minimally effective dose. In some embodiments, treatment is maintained at the lowest effective dose.

In another aspect, the invention provides, for example, expression levels of one or both of a biomarker, such as TARC or eotaxin-3, in a biological sample from a subject after administration of an IL-4R antagonist to the subject; or obtaining information about total serum levels of IgE; and that treatment should be continued if the expression levels of the biomarkers were decreased compared to levels prior to treatment with an IL-4R antagonist. A method of monitoring a subject's response to treatment with an IL-4R antagonist by providing instructions is featured, wherein the subject has moderate to severe asthma. In one embodiment, the biomarker is FeNO, and if FeNO levels are determined to decrease following administration of the antibody, instructions are provided to continue treatment with the IL-4R antagonist.

The present invention also provides for the treatment and/or prevention of asthma (e.g., eosinophilic asthma, moderate-to-severe eosinophilic asthma, etc.) or other indications or conditions disclosed herein. and an IL-4R antagonist disclosed herein for use in the manufacture of a medicament for treating any of

The present invention also provides for use in the treatment and/or prevention of asthma (e.g., eosinophilic asthma, moderate-to-severe eosinophilic asthma, etc.) or other indications disclosed herein. including IL-4R antagonists disclosed herein for the treatment and/or prevention of diseases or conditions.

The present invention includes pharmaceutical compositions comprising anti-IL4R antibody antagonists or antigen-binding fragments thereof for use in treating and/or preventing asthma and related conditions.

The invention also includes pharmaceutical compositions comprising anti-IL4R antibody antagonists or antigen-binding fragments thereof for use in reducing the incidence of one or more asthma exacerbations in a subject in need thereof. .

Additionally, the present invention includes pharmaceutical compositions comprising anti-IL4R antibody antagonists or antigen-binding fragments thereof for use in ameliorating one or more asthma-related parameters in a subject in need thereof.

The present invention provides a biomarker selected from the group consisting of thymic and activation-regulated chemokines (TARC), IgE, eotaxin-3, periostin, carcinoembryonic antigen (CEA), YKL-40, and exhaled nitric oxide (FeNO). pharmaceutical compositions comprising anti-IL4R antibody antagonists or antigen-binding fragments thereof for use in treating asthma and related conditions in patients with elevated levels of

The invention further comprises a pharmaceutical composition comprising an anti-IL4R antibody antagonist or antigen-binding fragment thereof for use in treating asthma or moderate-to-severe eosinophilic asthma in a subject in need thereof, wherein The treatment comprises testing the patient for the presence of a blood eosinophil level of at least 300 cells per microliter and/or a sputum eosinophil level of at least 3%, and blood eosinophils as described above Initiating/continuing administration of the pharmaceutical composition when globules and/or sputum eosinophil levels are found.

Other embodiments of the invention will become apparent from consideration of the detailed description below.

FIG. 1 is a graph showing a Kaplan-Meier plot of asthma exacerbation versus time in patients treated with placebo (open circles) compared to patients treated with anti-IL-4R antibody mAb1 (asterisks). The effect of treatment with anti-IL-4R antibody mAb1 persists for the time (including after 8 weeks) when patients are at higher risk of developing exacerbations due to steroid withdrawal. Vertical dashed lines indicate LABA withdrawals. FIG. 2 shows forced expiratory volume in liters (FEV1) in liters in placebo-treated patients (open triangles) compared to patients treated with anti-IL-4R antibody mAb1 (filled circles). Graph showing mean change from baseline. Vertical dashed lines indicate LABA withdrawals. FIG. 3 shows peak morning expiratory flow (AM PEF) in liters per minute in placebo-treated patients (open triangles) compared to patients treated with anti-IL-4R antibody mAb1 (filled circles). Graph showing mean change from baseline. FIG. 4 shows nighttime peak expiratory flow (PM PEF) in liters per minute in placebo-treated patients (open triangles) compared to patients treated with anti-IL-4R antibody mAb1 (filled circles). Graph showing mean change from baseline. FIG. 5 shows the mean change from baseline in inhaled albuterol use per day in placebo-treated patients (open triangles) compared to patients treated with anti-IL-4R antibody mAb1 (closed circles). It is a graph showing. FIG. 6. Mean change from baseline in 5-item Asthma Control Questionnaire (ACQ5) scores in placebo-treated patients (open triangles) compared to patients treated with anti-IL-4R antibody mAb1 (filled circles). is a graph showing Vertical dashed lines indicate LABA withdrawals. FIG. 7 is a graph showing the mean change from baseline in nocturnal awakenings per night in placebo-treated patients (open triangles) compared to patients treated with anti-IL-4R antibody mAb1 (filled circles). is. Vertical dashed lines indicate LABA withdrawals. Figure 8. Placebo-treated mITT population (filled circles) by visit at weeks 0, 1, 4, 8, and 12 compared to patients treated with anti-IL-4R antibody mAb1 (filled squares). FIG. 10 is a graph showing mean percentage change from baseline in TARC. FIG. Vertical dashed lines indicate LABA withdrawals. Figure 9. Placebo-treated mITT population (filled circles) compared to patients treated with anti-IL-4R antibody mAb1 (filled squares) by week 0, 1, 4, 8, and 12 visits. Figure 10 is a graph showing mean percentage change from baseline in eotaxin-3. Vertical dashed lines indicate LABA withdrawals. FIG. 10. By visit at Weeks 0, 1, 4, 8, and 12 in the mITT population treated with placebo (filled circles) compared to patients treated with anti-IL-4R antibody mAb1 (filled squares). FIG. 10 is a graph showing mean percentage change from baseline in total IgE. Vertical dashed lines indicate LABA withdrawals. Figure 11. Periostin by visits at weeks 0, 1, 4, 8, and 12 in placebo-treated mITT (filled circles) compared to patients treated with anti-IL-4R antibody mAb1 (filled squares). Graph showing the mean percentage change from baseline in . Figure 12. By visits at weeks 0, 1, 4, 8, and 12 in the mITT population treated with placebo (filled circles) compared to patients treated with anti-IL-4R antibody mAb1 (filled squares). FIG. 2 is a graph showing mean percentage change from baseline in carcinoembryonic antigen (CEA). Figure 13. By visits at Weeks 0, 1, 4, 8, and 12 in the mITT population treated with placebo (filled circles) compared to patients treated with anti-IL-4R antibody mAb1 (filled squares). Graph showing mean percentage change from baseline for YKL-40. FIG. 14. Weeks 0, 1, 2, 4, 6, 8, and 12 in the mITT population treated with placebo (filled circles) compared to patients treated with anti-IL-4R antibody mAb1 (filled squares). 1 is a graph showing the mean percentage change from baseline in blood eosinophils by 10-day visit. FIG. 15. Baseline by visits 0, 4, 8, and 12 in the placebo-treated mITT population (filled circles) compared to patients treated with anti-IL-4R antibody mAb1 (filled squares). 1 is a graph showing the average percentage change in exhaled nitric oxide (NO) levels from. Vertical dashed lines indicate LABA withdrawals. FIG. 16 shows exhaled nitric oxide (FeNO) in the placebo-treated mITT population (open circles and full line) compared to patients treated with the anti-IL-4R antibody mAb1 (plus sign and dotted line). PPB) is a scatter plot of change in FEV1 (L) from baseline as a percentage of baseline at week 12. FIG. FIG. 17 shows the baseline FeNO (PPB) versus baseline at week 12 in the placebo-treated mITT population (open circles and solid line) compared to patients treated with anti-IL-4R antibody mAb1 (plus sign and dotted line). Scatter plot of change in AM-PEF (L/min) from line. FIG. 18 shows baseline FeNO (PPB) vs. baseline FeNO (PPB) in placebo-treated mITT population (open circles and solid line) compared to patients treated with anti-IL-4R antibody mAb1 (plus sign and dotted line) at week 12. Scatter plot of change in PM-PEF (L/min) from line. FIG. 19 shows blood eosinophil counts (GIGA/L) in placebo-treated mITT population (open circles and solid line) compared to patients treated with anti-IL-4R antibody mAb1 (plus sign and dotted line). Scatter plot of change in FEV1 from baseline at week 12 (L). FIG. 20 shows blood eosinophil counts (GIGA/L) in placebo-treated mITT population (open circles and solid line) compared to patients treated with anti-IL-4R antibody mAb1 (plus sign and dotted line). FIG. 4 is a scatter plot of ACQ change from baseline at Week 12. FIG. FIG. 21 shows blood eosinophil counts (GIGA/L) in placebo-treated mITT population (open circles and solid line) compared to patients treated with anti-IL-4R antibody mAb1 (plus sign and dotted line). FIG. 10 is a scatter plot of change in daily albuterol/lev-albuterol use from baseline at week 12 vs. FIG. 22. Week 12 baseline-to-baseline change for baseline periostin in placebo-treated mITT population (open circles and solid line) compared to patients treated with anti-IL-4R antibody mAb1 (plus sign and dotted line). FIG. 4 is a scatter plot of changes in ACQ; FIG. 23. Week 12 baseline to YKL-40 in placebo-treated mITT population (open circles and solid line) compared to patients treated with anti-IL-4R antibody mAb1 (plus sign and dotted line). FIG. 4 is a scatter plot of changes in ACQ; FIG. 24 is a schematic representation of timing and dosing regimens for treatment of asthma patients. Figure 25 has persistent moderate-to-severe eosinophilic asthma partially controlled/uncontrolled with inhaled corticosteroid (ICS) and long-acting beta-2 agonist (LABA) treatment. A schematic diagram describing the patient disposition of a randomized, placebo-controlled, double-blind, parallel-group study conducted with patients subcutaneously administered either 300 mg mAb1 or placebo once weekly for 12 weeks. be. Figures 26A and 26B are scatter plots of morning (A) and evening (B) asthma symptoms measured over 12 weeks after administration of placebo (open triangles) or mAb1 (filled circles). FIG. 27 shows humanized IL-4/IL-4R following house dust mite (HDM) exposure and treatment with either anti-IL-4R antibody or IL-13Ra2-Fc decoy receptor molecule, or mock treatment. FIG. 4 is a graph showing serum IgE levels in mice (IL-4 ^hu/hu IL-4Rα ^hu/hu ). Measurements were made on samples taken at the end of the study on Day 40 (24 hours prior to the first dose of treatment) and Day 85. FIG. 28 shows results in wild-type (Balb/c) mice after house dust mite (HDM) exposure and isotype control, treatment with either anti-IL-4R antibody or IL-13Ra2-Fc decoy receptor molecule, or sham treatment. Figure 2 is a graph showing serum IgE levels; FIG. 29 is a graph showing lung collagen content (expressed in μg/lobe) of humanized IL-4/IL-4R mice after HDM exposure and the indicated treatments. FIG. 30 is a graph showing collagen content (expressed in μg/lobe of lung) in lungs of wild-type mice after HDM exposure and the indicated treatments. Figure 31A is a graph showing levels of eosinophils and neutrophils in humanized IL-4/IL-4R mice after HDM exposure and the indicated treatments, and Figure 31B shows HDM exposure and the indicated treatments. FIG. 4 is a graph showing levels of resident and inflammatory dendritic cells in humanized IL-4/IL-4R mice after treatment.

DETAILED DESCRIPTION Before describing this invention, it is to be understood that this invention is not limited to the particular methods and experimental conditions described, as such methods and conditions may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting. This is because the scope of the present invention is limited only by the appended claims.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

The term "about," as used herein with respect to a particular stated numerical value, means that the numerical value may vary from the stated value by no more than 1%. For example, the phrase "about 100" as used herein includes 99 and 101 and all values between (eg, 99.1, 99.2, 99.3, 99.4, etc.).

As used herein, the terms "treat," "treating," etc., alleviate the symptoms of, or eliminate, either temporarily or permanently, the cause of the symptoms of the named disorder or condition. or to prevent or delay the appearance of symptoms.

Although any methods and materials similar or equivalent to those described herein can be used in the practice of the invention, the preferred methods and materials are now described. All publications mentioned herein are hereby incorporated by reference in their entirety.

Methods for Reducing the Incidence of Asthma Exacerbations The present invention includes methods for reducing the incidence of asthma exacerbations in a subject in need thereof, which methods comprise interleukin-4 receptor (IL- 4R) administering to the subject a pharmaceutical composition comprising the antagonist. As used herein, the phrase "asthma exacerbation" means an increase in the severity and/or frequency and/or duration of one or more symptoms or signs of asthma. An "asthma exacerbation" is also any condition in a subject's respiratory health that requires and/or is treatable by an asthma intervention (e.g., steroid treatment, inhaled corticosteroid treatment, hospitalization, etc.). including the deterioration of According to certain embodiments of the invention, an asthma exacerbation is defined as one or more of the following: (a) morning peak expiratory flow (“AM PEF”, herein (b) 6 or more doses of albuterol or lev-albuterol (compared to baseline) in 24 hours on 2 consecutive days; and (c) (i) systemic (oral and/or parenteral) steroid treatment, or (ii) increase in inhaled corticosteroids to at least four times baseline levels, or (iii) hospitalization. exacerbation of asthma (eg, as determined by a physician or other medical practitioner) requiring

In certain instances, an asthma exacerbation can be classified as a "severe asthma exacerbation." A "severe asthma exacerbation" is an event requiring immediate intervention in the form of treatment with either systemic corticosteroids or inhaled corticosteroids at doses four times or greater than those taken prior to the event. means. Thus, the general expression "asthma exacerbation" includes and encompasses the more specific subcategory of "severe asthma exacerbation." Accordingly, the present invention includes methods for reducing the incidence of severe asthma exacerbations in patients in need thereof.

A "reduced incidence" of asthma exacerbations means that a subject administered a pharmaceutical composition of the invention experiences fewer asthma exacerbations after treatment than before treatment (i.e., at least one less exacerbation), or It means not experiencing an asthma exacerbation for at least 4 weeks (eg 4, 6, 8, 12, 14 or more weeks) after initiation of treatment with a pharmaceutical composition of the invention. Alternatively, a reduction in the "incidence of asthma exacerbations" means that a subject is more likely to experience an asthma exacerbation after administration of a composition of the invention compared to a subject not administered a pharmaceutical composition of the invention. , means a reduction of at least 10% (eg, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% or more).

Methods for Improving Asthma-Related Parameters The invention also includes methods for improving one or more asthma-related parameters in a subject in need thereof, wherein the methods comprise interleukin-4 administering to the subject a pharmaceutical composition comprising a receptor (IL-4R) antagonist. For purposes of the present invention, a reduction in the incidence of asthma exacerbations (as described above) may be correlated with improvement in one or more asthma-related parameters; is not observed in

Examples of "asthma-related parameters" include: (a) forced expiratory volume in one second (FEV1); (b) peak expiratory flow (PEF), including morning PEF (AM PEF) and evening PEF (PM PEF); (d) 5-item Asthma Control Questionnaire (ACQ5) score; (d) nocturnal awakening; and (e) 22-item sinus assessment test (SNOT-22). ) score. "Improvement in asthma-related parameters" includes an increase from baseline in one or more of FEV1, AM PEF or PM PEF, and/or one or more daily albuterol/lev-albuterol use, ACQ5 score, Mean nocturnal alertness or decrease from baseline in SNOT-22 score. The term "baseline" as used herein with respect to asthma-related parameters means the numerical values of asthma-related parameters for a patient prior to administration of the pharmaceutical composition of the invention or at the time of administration of the pharmaceutical composition of the invention. do.

To determine whether an asthma-related parameter has been "improved," the parameter is quantified at baseline and at time points after administration of the pharmaceutical composition of the invention. For example, asthma-related parameters may be measured after 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days after initial treatment with a pharmaceutical composition of the invention. , 12 days, 14 days, or 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 13 weeks after, 14 weeks, 15 weeks, 16 weeks, 17 weeks, 18 weeks, 19 weeks, 20 weeks, 21 weeks, 22 weeks, 23 weeks, 24 weeks or more can be measured. The difference between the value of the parameter at a specific time point after initiation of treatment and the value of the parameter at baseline is used to "improve" an asthma-related parameter (e.g., depending on whether there was an increase or decrease, as the case may be.

The term "acquire" or "acquiring" as used herein means "directly obtaining" or "indirectly obtaining" a physical entity or a value such as an asthma-related parameter. refers to possession of a physical entity or acquisition of a value, such as a number. "Directly obtaining" means performing a process (eg, performing a synthetic or analytical method) to obtain a physical entity or value. "Indirectly obtained" refers to receiving a physical entity or value from another party or source (eg, a third party laboratory that directly obtained the physical entity or value). Obtaining physical substance directly involves performing a process that involves a physical change in a physical substance, eg, a starting material. Exemplary changes include making physical entities from two or more starting materials, shearing or fragmenting materials, separating or purifying materials, separating or purifying materials from two or more separate entities. are combined into a mixture, and chemical reactions involving breaking or forming covalent or non-covalent bonds are performed. Obtaining a value directly includes performing a process that involves a physical change in a sample or another substance, e.g., an analytical process that involves a physical change in a substance, e.g. (sometimes referred to as “analyses”).

Indirectly obtained information may be provided in the form of reports obtained, for example, in paper or electronic form, such as from an online database or application (“App”). This report or information may be provided by, for example, a health care agency, such as a hospital or clinic; or a health care provider, such as a doctor or nurse.

Forced expiratory volume in 1 second (FEV1) . According to certain embodiments of the invention, administration of an IL-4R antagonist to a patient results in an increase from baseline in forced expiratory volume in one second (FEV1). Methods for measuring FEV1 are known in the art. For example, a spirometer meeting the 2005 American Thoracic Society (ATS)/European Respiratory Society (ERS) recommendations can be used to measure FEV1 in a patient. ATS/ERS Standardization of Spirometry can be used as a guideline. Spirometry is generally performed between 6 and 10 AM after at least 6 hours of albuterol retention. Pulmonary function tests are generally performed in a sitting position and the highest measurement is recorded as FEV1 (liters).

The present invention includes therapeutic methods that result in an increase in FEV1 from baseline of at least 0.05 L 12 weeks after initiation of treatment with a pharmaceutical composition comprising an anti-IL-4R antagonist. For example, according to the present invention, administration of an IL-4R antagonist to a subject in need thereof at week 12 is about 0.05L, 0.10L, 0.12L, 0.14L, 0.16L , 0.18L, 0.20L, 0.22L, 0.24L, 0.26L, 0.28L, 0.30L, 0.32L, 0.34L, 0.36L, 0.38L, 0.40L, 0 resulting in an increase in FEV1 from baseline of 0.42L, 0.44L, 0.46L, 0.48L, 0.50L, or more.

Morning and evening peak expiratory flow (AM PEF and PM PEF) . According to a particular embodiment of the invention, the administration of an IL-4R antagonist to a patient reduces morning (AM) and/or evening (PM) peak expiratory flow (AM PEF and/or PM PEF) from baseline to resulting in an increase in Methods for measuring PEF are known in the art. For example, according to one method for measuring PEF, patients require morning (AM) and evening (PM) PEF (as well as daily albuterol use, morning and evening asthma symptom scores, and rescue medication). You will be issued an electronic PEF instrument to read the number of nocturnal awakenings due to asthma symptoms. The patient is instructed on the use of the device and written instructions on the use of the electronic PEF meter are provided to the patient. Additionally, the medical professional may instruct the patient on how to record the relevant variables on the electronic PEF instrument. AM PEF is performed within 15 minutes after waking up (between 6 am and 10 am), generally prior to any albuterol intake. PM PEF is generally done in the evening (between 6pm and 10pm) before taking any albuterol. Subjects should try to withhold albuterol for at least 6 hours before measuring their PEF. Three PEF attempts are performed by the patient and all three values are recorded by an electronic PEF instrument. Usually the highest value is used for evaluation. Baseline AM PEF may be calculated as the mean AM measurements recorded during the 7 days prior to administration of the first dose of the pharmaceutical composition comprising the IL-4R antagonist, and baseline PM PEF comprising the IL-4R antagonist. May be calculated as the mean PM measurements recorded during the 7 days prior to administration of the first dose of the pharmaceutical composition.

The present invention provides a method of treatment that results in an increase in AM PEF and/or PM PEF from baseline of at least 1.0 L/min 12 weeks after initiation of treatment with a pharmaceutical composition comprising an anti-IL-4R antagonist. include. For example, according to the present invention, administration of an IL-4R antagonist to a subject in need thereof is at about 0.5 L/min, 1.0 L/min, 1.5 L/min, 2 .0 L/min, 2.5 L/min, 3.0 L/min, 3.5 L/min, 4.0 L/min, 4.5 L/min, 5.0 L/min, 5.5 L/min, 6.0 L /min, 6.5 L/min, 7.0 L/min, 7.5 L/min, 8.0 L/min, 8.5 L/min, 9.0 L/min, 9.5 L/min, 10.0 L/min , 10.5 L/min, 11.0 L/min, 12.0 L/min, 15 L/min, 20 L/min, or more resulting in an increase in PEF from baseline.

Use of albuterol/levalbuterol . According to certain embodiments of the invention, administration of an IL-4R antagonist to a patient results in a decrease from baseline in daily albuterol or lev-albuterol use. The number of albuterol/lev-albuterol inhalations may be recorded daily by the patient on a diary, PEF instrument, or other recording device. During treatment with the pharmaceutical composition of the present invention, the use of albuterol/levalbuterol may not be regular or prophylactic, typically as needed for symptoms. The baseline number of albuterol/lev-albuterol inhalations/day may be calculated based on the average of the 7 days prior to administration of the first dose of the pharmaceutical composition comprising the IL-4R antagonist.

Treatment methods that result in a reduction in albuterol/lev-albuterol usage from baseline of at least 0.25 puffs per day 12 weeks after initiation of treatment with a pharmaceutical composition comprising an anti-IL-4R antagonist. For example, according to the present invention, administration of an IL-4R antagonist to a subject in need thereof at week 12 is about 0.25 puffs per day, 0.50 puffs per day, 0.75 puffs, 1.00 puffs per day, 1.25 puffs per day, 1.5 puffs per day, 1.75 puffs per day, 2.00 puffs per day, 2.25 puffs per day, 1 Resulting in a reduction in albuterol/lev-albuterol usage from baseline of 2.5 puffs per day, 2.75 puffs per day, 3.00 puffs per day or more.

Five-item Asthma Control Questionnaire (ACQ) scores . According to certain embodiments of the invention, administration of an IL-4R antagonist to a patient results in a decrease from baseline in the 5-item Asthma Control Questionnaire (ACQ5) score. ACQ5 is a validated questionnaire for assessing asthma management.

Treatment methods that result in a reduction in ACQ5 score from baseline of at least 0.10 points 12 weeks after initiation of treatment with a pharmaceutical composition comprising an anti-IL-4R antagonist. For example, according to the present invention, administration of an IL-4R antagonist to a subject in need thereof will be about 0.10 points, 0.15 points, 0.20 points, 0.25 points at week 12. , 0.30 points, 0.35 points, 0.40 points, 0.45 points, 0.50 points, 0.55 points, 0.60 points, 0.65 points, 0.70 points, 0.75 points , 0.80 points, 0.85 points or greater reduction in ACQ score from baseline.

Awakening at night . According to certain embodiments of the invention, administration of an IL-4R antagonist to a patient results in a decrease from baseline in the average number of nocturnal awakenings.

The present invention provides a method of treatment that results in a reduction in the average number of nocturnal awakenings from baseline of at least about 0.10 per night 12 weeks after initiation of treatment with a pharmaceutical composition comprising an anti-IL-4R antagonist. include. For example, according to the present invention, administration of an IL-4R antagonist to a subject in need thereof will be approximately 0.10 times per night, 0.15 times per night, 0.15 times per night at week 12, 0.20 times per night 0.25 times per night 0.30 times per night 0.35 per night 0.40 per night 0.45 per night 0.25 per night 50 times per night 0.55 times per night 0.60 per night 0.65 per night 0.70 per night 0.75 per night 0.80 per night , 0.85 nightly, 0.90 nightly, 0.95 nightly, 1.0 nightly, 2.0 nightly or more nocturnal awakenings from baseline result in a decrease in the average number of

22-item sinus score test (SNOT-22) score . According to certain embodiments of the invention, administration of an IL-4R antagonist to a patient results in a reduction from baseline in the 22-item sinus score test (SNOT-22). SNOT-22 is a valid questionnaire to assess the impact of chronic sinusitis on quality of life (Hopkins et al 2009, Clin. Otolaryngol. 34:447-454).

The present invention includes therapeutic methods that result in a reduction in SNOT-22 score from baseline of at least 1 point 12 weeks after initiation of treatment with a pharmaceutical composition comprising an anti-IL-4R antagonist. For example, according to the present invention, administration of an IL-4R antagonist to a subject in need thereof reduces the SNOT-22 score from baseline by about 1, 2, 3, 4, 5, 1, 2, 3, 4, 5 at week 12. Resulting in a reduction of 6, 7, 8, 9, 10, 11, 12, 13 points or more.

Methods for Treating Asthma According to certain embodiments, the present invention provides methods for treating asthma (including, for example, eosinophilic asthma) in a subject in need thereof, comprising: The method comprises administering to the subject a pharmaceutical composition comprising an interleukin-4 receptor (IL-4R) antagonist. In certain embodiments, the methods of the invention are useful for treating moderate to severe eosinophilic asthma (e.g., persistent moderate to severe eosinophilic asthma) in a subject. .

According to the present invention, a subject is moderate to severe when he or she exhibits a blood eosinophil level of at least 300 cells per microliter and/or a sputum eosinophil level of at least 3%. identified as having eosinophilic asthma. Any method known and available in the art for measuring blood and/or sputum eosinophil levels can be used in the context of the present invention if the subject has moderate to severe eosinophilic asthma. and therefore identified as suitable subjects for the therapeutic methods of the present invention.

According to a related aspect of the invention, selecting patients exhibiting a blood eosinophil level of at least 300 cells per microliter and/or a sputum eosinophil level of at least 3%; and (b) IL Methods are provided for treating asthma comprising administering to the patient a pharmaceutical composition comprising a -4R antagonist.

In another aspect, a method for reducing or eliminating an asthma patient's dependence on inhaled corticosteroids (ICS) and/or long-acting beta-agonists (LABAs) during treatment of moderate to severe asthma. is provided. In certain embodiments, the method comprises: selecting a patient with moderate to severe asthma uncontrolled or partially controlled with background asthma therapy; administering an IL-4R antibody to the patient during the initial treatment while maintaining the patient's background asthma therapy during the initial treatment period; and continuing administration of the IL-4R antagonist over the subsequent treatment period. Including gradually reducing the dosage of one or more components of the background therapy. "Background therapy" refers to standard or conventional therapeutic agents known in the art used to treat asthma. In certain embodiments, the background therapy comprises ICS, LABA, or a combination of both. In some embodiments, dosages of ICS and/or LABA are eliminated or completely withdrawn during the initial treatment period. For example, a LABA such as salmeterol or formoterol is administered during an initial treatment period and completely stopped or withdrawn during subsequent treatment periods.

An example of a treatment regimen for a patient with moderate to severe asthma is shown in Figure 24, wherein an IL-4R antagonist is administered to a patient with moderate to severe asthma. LABA and ICS are administered to patients as background therapy during the initial treatment period (also referred to as the "stabilization period"). During the follow-up treatment period (also called the "washout period"), administration of LABA is stopped, ie LABA is withdrawn or eliminated. ICS is gradually decreased over the subsequent treatment period until it is eliminated.

In a related aspect, methods are provided for treating asthma that include adjunctive therapies to background therapies while withholding systemic background therapies. In certain embodiments, the IL-4R antagonist is in background therapy for a specified period of time (e.g., 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 5 months, 12 months, 18 months, 24 months or longer) (also called "stable phase") as an adjunctive therapy to asthma patients. In some embodiments, the background therapy comprises ICS and/or LABA. The stable period is followed by a background treatment withdrawal period, in which one or more components that make up the background treatment are withdrawn, reduced or eliminated, but not the adjunctive treatment. continues. In some embodiments, background therapy may be reduced by about 5%, about 10%, about 20%, about 30%, about 40%, about 50% or more during the drug holiday. The drug holiday may last 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks or longer. In preferred embodiments, the background treatment may be reduced by about 5% during the washout period, and the washout period is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks. , 9 weeks, 10 weeks, 11 weeks, 12 weeks or longer. In preferred embodiments, the background treatment may be reduced by about 10% during the washout period, and the washout period is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks. , 9 weeks, 10 weeks, 11 weeks, 12 weeks or longer. In preferred embodiments, the background treatment may be reduced by about 20% during the washout period, and the washout period is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks. , 9 weeks, 10 weeks, 11 weeks, 12 weeks or longer. In preferred embodiments, the background treatment may be reduced by about 30% during the washout period, and the washout period is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks. , 9 weeks, 10 weeks, 11 weeks, 12 weeks or longer. In preferred embodiments, the background treatment can be reduced by about 40% during the washout period, and the washout period is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks. , 9 weeks, 10 weeks, 11 weeks, 12 weeks or longer. In preferred embodiments, the background treatment may be reduced by about 50% or more during the washout period, wherein the washout period is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, It can last 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks or longer.

In some other embodiments, the present invention provides asthma-related conditions or complications, such as chronic sinusitis, allergic rhinitis, allergic fungal rhinosinusitis, allergic bronchopulmonary aspergillosis, integrative airway disease. Included are methods of treating or alleviating unified airline disease, Churg-Strauss syndrome, vasculitis, chronic obstructive pulmonary disease (COPD), and exercise-induced bronchospasm.

The invention also includes a method of treating persistent asthma. As used herein, the term "persistent asthma" means having symptoms at least once a week during the day and/or at night, the symptoms lasting hours to days. In certain alternative embodiments, persistent asthma is defined as "mildly persistent" (e.g., symptoms severe enough to interfere with daily activities or sleep and more than twice a week but less than daily, and /or pulmonary function is normal or reversible with inhaled bronchodilators), "moderately persistent" (e.g., sleep is disturbed at least weekly and/or pulmonary function is moderately abnormal) or "severely persistent" (e.g., when symptoms persist despite correct use of approved medications and/or pulmonary function is severely affected). be.

Interleukin-4 Receptor Antagonists The methods of the invention involve administering to a subject in need thereof a therapeutic composition comprising an interleukin-4 receptor (IL-4R) antagonist. As used herein, an "IL-4R antagonist" binds to or interacts with IL-4R and, when IL-4R is expressed on a cell in vitro or in vivo, an IL-4R antagonist. Any agent that inhibits the normal biological signaling function of the 4Rs. Non-limiting examples of categories of IL-4R antagonists include small molecule IL-4R antagonists, anti-IL-4R aptamers, peptide-based IL-4R antagonists (eg, "peptibody" molecules), and human IL-4R antagonists. Antibodies or antigen-binding fragments of antibodies that specifically bind to -4R are included.

The term "human IL4R" (hIL-4R) refers to a human cytokine receptor that specifically binds interleukin-4 (IL-4), such as IL-4Rα (SEQ ID NO:274).

The term "antibody" refers to an immunoglobulin molecule comprising four polypeptide chains, two heavy (H) chains and two light (L) chains interconnected by disulfide bonds, as well as multimers thereof (e.g., IgM). point to Each heavy chain comprises a heavy chain variable region (abbreviated herein as HCVR or _VH ) and a heavy chain constant region. The heavy chain constant region comprises three domains, C _H 1, C _H 2, and C _H 3. Each light chain comprises a light chain variable region (abbreviated herein as LCVR or _VL ) and a light chain constant region. The light chain constant region contains one domain (C _L 1). The V _H and V _L regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each V _H and V _L is composed of three CDRs and four FRs arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. In different embodiments, the FRs of the anti-IL-4R antibody (or antigen-binding fragment thereof) can be identical to human germline sequences, or can be naturally or artificially altered. An amino acid consensus sequence can be defined based on a side-by-side analysis of two or more CDRs.

The term "antibody" also includes antigen-binding fragments of full antibody molecules. As used herein, the terms "antigen-binding portion" of an antibody, "antigen-binding fragment" of an antibody, etc., refer to any naturally occurring or enzymatic antibody that specifically binds to an antigen to form a complex. or synthetic or genetically engineered polypeptides or glycoproteins. Antigen-binding fragments of antibodies are produced using any suitable standard technique, such as proteolytic digestion, or recombinant genetic engineering techniques, including the manipulation and expression of the DNA encoding the variable facets and optionally the constant domains of the antibodies. As such, it can be derived, for example, from a complete antibody molecule. Such DNA is known and/or readily available, eg, from commercial sources, DNA libraries (including, eg, phage-antibody libraries), or can be synthesized. The DNA may be modified, for example, to place one or more variable and/or constant domains in an appropriate configuration, or to introduce codons, create cysteine residues, or alter, add or delete amino acids. For example, it can be sequenced and manipulated chemically or by using molecular biology techniques.

(ii) F(ab')2 fragment; (iii) Fd fragment; (iv) Fv fragment; (v) single chain Fv (scFv). (vi) a dAb fragment; and (vii) a minimal recognition unit (e.g., an isolated complementarity determining region (CDR), such as a CDR3 peptide) consisting of amino acid residues that mimics the hypervariable regions of an antibody, or a constraint. and constrained FR3-CDR3-FR4 peptides. Other engineered molecules such as domain specific antibodies, single domain antibodies, domain deleted antibodies, chimeric antibodies, CDR grafted antibodies, bispecific antibodies, triabodies, tetraspecific antibodies (tetrabodies), minibodies, nanobodies (e.g., monovalent nanobodies, bivalent nanobodies, etc.), small modular immunopharmaceuticals (SMIPs), and shark variable IgNAR domains are also expressed as "antigen-binding fragments." contained within.

An antigen-binding fragment of an antibody typically comprises at least one variable domain. A variable domain can be of any size or amino acid composition and generally includes at least one CDR flanking or in-frame with one or more framework sequences. . In antigen-binding fragments having a _VH domain associated with a _VL domain, the _VH and _VL domains may be positioned relative to each other in any suitable arrangement. For example, the variable region can be dimeric, V _H -V _H , V _H -V _L or V _L
-V _L dimers. Alternatively, the antigen-binding fragment of an antibody may be a monomeric _VH or _VL
May contain domains.

In certain embodiments, an antigen-binding fragment of an antibody may comprise at least one variable domain covalently linked to at least one constant domain. Non-limiting, exemplary configurations of variable and constant domains that may be found in antigen-binding fragments of antibodies of the invention include: (i) V _H -C _H 1; (ii) V _H -C _H 2; (iii) V _H -C _H 3; (iv) V _H -C _H 1-C _H 2; (v) V _H -C _H 1-C _H 2-C _H 3; (vi) V _H -C _H 2-CH3; (vii) _VH - _CL ; (viii) _VL - _CH1 ; (ix) _VL - _CH2 ; (x) _VL - _CH3 ; (xi _{) VL} (xii) V _L -C _H 1-C _H 2- _{C H 3; (xiii) V L -C H 2-C H 3 ; and ( xiv} ) V _L -C _L are mentioned. In any of the configurations of the variable and constant domains, including any of the exemplary configurations listed above, the variable and constant domains are directly linked to each other, or by full-length or partial hinge or linker regions. can be concatenated. The hinge regions are at least two (eg, 5, 10, 15, 20, 40) that create flexible or semi-flexible linkages between adjacent variable and/or constant domains in a single polypeptide molecule. , 60 or more) amino acids, preferably the hinge region may consist of between 2-60 amino acids, preferably 5-50, or preferably 10-40 amino acids. Further, antigen-binding fragments of the antibodies of the invention can be combined with each other and/or with one or more monomeric V
comprising homodimers or heterodimers (or other multimers) of any of the above-listed variable and constant domain configurations non-covalently linked (e.g., by disulfide bonds) to _H or V _L domains obtain.

As with full antibody molecules, antigen-binding fragments can be monospecific or multispecific (eg, bispecific). A multispecific antigen-binding fragment of an antibody typically comprises at least two different variable domains, wherein each variable domain can specifically bind to a separate antigen or different epitopes on the same antigen. Any multispecific antibody format can be adapted for use in the context of antigen-binding fragments of the antibodies of the invention using routine techniques available in the art.

The constant region of an antibody is important in the antibody's ability to fix complement and mediate cell-dependent cytotoxicity. Thus, the isotype of the antibody can be selected based on whether it is desirable for the antibody to mediate cytotoxicity.

The term "human antibody" includes antibodies having variable and constant regions derived from human germline immunoglobulin sequences. Nonetheless, the human antibodies featured in the present invention may have amino acid residues not encoded by human germline immunoglobulin sequences, e.g. It may contain mutations introduced by somatic mutation, but the term "human antibody" refers to antibodies in which CDR sequences derived from the germline of another mammalian species, such as mouse, have been grafted onto human framework sequences. Not included.

The term "recombinant human antibody" refers to any human antibody produced, expressed, generated or isolated by recombinant means, e.g., an antibody expressed using a recombinant expression vector transfected into a host cell (hereinafter ), antibodies isolated from recombinant combinatorial human antibody libraries (further described below), antibodies isolated from animals (e.g. mice) transgenic for human immunoglobulin genes (e.g. , Taylor
et al. (1992) Nucl. Acids Res. 20:6287-6295) or by any other means including splicing of the human immunoglobulin gene sequences into other DNA sequences. Such recombinant human antibodies have variable and constant regions derived from human germline immunoglobulin sequences. However, in certain embodiments, such recombinant human antibodies are subjected to in vitro mutagenesis (or in vivo somatic mutagenesis if animals transgenic for human Ig sequences are used), thus The amino acid sequences of the _VH and _VL regions of the antibody are derived from and related to human _{germline VH and VL sequences} , but in vivo within the human antibody germline repertoire are: may not occur naturally.

Human antibodies can exist in two forms related to hinge heterogeneity. In one form, the immunoglobulin molecule comprises a stable four-chain construction of about 150-160 kDa, wherein the dimers are held together by interchain heavy chain disulfide bonds. In a second form, the dimer is a molecule of approximately 75-80 kDa that is not linked via interchain disulfide bonds and is composed of covalently coupled light and heavy chains (half antibodies). is formed. These forms were very difficult to separate even after affinity purification.

The frequency of occurrence of the second form in various intact IgG isotypes is due to, but not limited to, structural differences associated with antibody hinge region isotypes. A single amino acid substitution in the hinge region of the human IgG4 hinge can significantly reduce the appearance of the second form to levels typically observed using the human IgG1 hinge (Angal et al.
al. (1993) Molecular Immunology 30:105).
The invention encompasses antibodies with one or more mutations in the hinge, C _H 2, or C _H 3 regions, which may be desirable in manufacturing, for example, to improve the yield of the desired antibody form. do not have.

"Isolated antibody" means an antibody that has been identified and separated and/or recovered from at least one component of its natural environment. For example, an antibody that has been separated or removed from at least one component of an organism or from a tissue or cell in which it naturally occurs or is produced is an "isolated antibody" for the purposes of the present invention. ”. Isolated antibody also includes the antibody in situ within recombinant cells. An isolated antibody is an antibody that has undergone at least one purification or isolation step. According to certain embodiments, an isolated antibody may be substantially free of other cellular material and/or chemicals.

The term "specifically binds" or the like means that an antibody or antigen-binding fragment thereof forms a relatively stable complex with an antigen under physiological conditions. Methods for determining whether an antibody specifically binds an antigen are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, and the like. For example, an antibody that "specifically binds" to IL-4R as used in the context of the present invention will bind IL-4R or a portion thereof to less than about 1000 nM, less than about 500 nM, less than about 500 nM, less than about less than 300 nM, less than about 200 nM, less than about 100 nM, less than about 90 nM, less than about 80 nM, less than about 70 nM, less than about 60 nM, less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 5 nM , less than about 4 nM, less than about 3 nM, less than about 2 nM, less than about 1 nM, or less than about 0.5 _nM . However, human IL-4
An isolated antibody that specifically binds R may have cross-reactivity to other antigens, such as IL-4R molecules from other (non-human) species.

An anti-IL-4R antibody useful for the methods of the invention will have one sequence in the framework and/or CDR regions of the heavy and light chain variable domains compared to the corresponding germline sequences from which the antibody is derived. one or more amino acid substitutions, insertions, and/or deletions (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 substitutions and/or 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 insertions and/or 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 deletions). Such mutations can be readily ascertained by comparing the amino acid sequences disclosed herein to germline sequences available, for example, from public antibody sequence databases. The invention includes methods comprising the use of antibodies and antigen-binding fragments thereof derived from any of the amino acid sequences disclosed herein, wherein one or more frameworks and/or one or more (e.g. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 for tetrameric antibodies or 1, 2, 3, 4, 5 for HCVR and LCVR of antibodies) or 6) one or more amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids) within the CDR regions of the antibody is derived from the germline The corresponding residue of the sequence, or the corresponding residue of another human germline sequence, or the corresponding germline residue is mutated to a conservative amino acid substitution (such sequence changes are collectively referred to herein as called “germline mutation”). One skilled in the art can readily produce large numbers of antibodies and antigen-binding fragments containing one or more individual germline mutations or combinations thereof starting from the heavy and light chain variable region sequences disclosed herein. can do. In certain embodiments, all framework and/or CDR residues within the V _H and/or V _L domains are backmutated to those residues found in the germline sequence from which the antibody was derived. In other embodiments, only certain residues, e.g., only mutated residues found within the first eight amino acids of FR1 or within the last eight residues of FR4, or only those found within CDR1, CDR2 or CDR3. Only the mutated residues that are mutated are backmutated to the original germline sequence. In other embodiments, one or more of the framework and/or CDR residues are corresponding residues in a different germline sequence (i.e., a germline sequence that differs from the germline sequence from which the antibody was originally derived). mutated into. Furthermore, antibodies of the invention may contain any combination of two or more germline mutations in the framework and/or CDR regions, e.g., where a particular individual residue is a particular germline While mutated to the corresponding residue of the sequence, certain other residues that differ from the original germline sequence are either maintained or mutated to the corresponding residue of the different germline sequence. Once obtained, antibodies and antigen-binding fragments comprising one or more germline mutations have improved binding specificity, increased binding affinity, improved or enhanced antagonistic or agonistic biological properties. It can (optionally) be readily tested for one or more desired properties such as reduced immunogenicity and the like. The use of antibodies and antigen-binding fragments obtained in this general fashion is encompassed within the invention.

The invention also contemplates the use of anti-IL-4R antibodies comprising variants of any of the HCVR, LCVR and/or CDR amino acid sequences disclosed herein having one or more conservative substitutions. Including how to include. For example, the present invention provides, for example, 10 or less, 8 or less, 6 or less, 4 or more, compared to any of the HCVR, LCVR, and/or CDR amino acid sequences disclosed herein. Includes use of anti-IL-4R antibodies having HCVR, LCVR, and/or CDR amino acid sequences containing conservative amino acid substitutions such as:

The term "surface plasmon resonance" refers to the analysis of real-time interactions by detecting changes in protein concentration within a biosensor matrix, for example using the BIAcore ^™ system (Biacore Life Sciences division of GE Healthcare, Piscataway, NJ). refers to an optical phenomenon that enables

The term "K _D " refers to the equilibrium dissociation constant for a particular antibody-antigen interaction.

The term "epitope" refers to antigenic determinants that interact with specific antigen-binding sites in the variable regions of antibody molecules known as paratopes. A single antigen can have more than one epitope. Different antibodies may therefore bind to different regions of the antigen and have different biological effects. Epitopes can be either conformational or linear. Conformational epitopes arise from spatially adjacent amino acids from different segments of a linear polypeptide chain. A linear epitope is one produced by adjacent amino acid residues in a polypeptide chain. In certain circumstances, epitopes may include sugar, phosphoryl, or sulfonyl moieties on the antigen.

Production of Human Antibodies Methods for producing human antibodies in transgenic mice are known in the art. Any such method can be used in the context of the present invention to generate human antibodies that specifically bind human IL-4R.

VELOCIMMUNE ^™ technology (see, e.g., US Pat. No. 6,596,541, Regeneron Pharmaceuticals) or any other known method for generating monoclonal antibodies, having a human variable region and a mouse constant region, A high affinity chimeric antibody to IL-4R is first isolated. VELOCIMMUNE ^(R) technology comprises human heavy chains operably linked to endogenous mouse constant region loci such that mice produce antibodies containing human variable regions and mouse constant regions in response to antigenic stimulation. and the generation of transgenic mice whose genome contains the light chain variable region. The DNA encoding the heavy and light chain variable regions of the antibody is isolated and operably linked to DNA encoding the human heavy and light chain constant regions. The DNA is then expressed in cells capable of expressing fully human antibodies.

Generally, ^{VELOCIMMUNE®} mice are exposed to the antigen of interest, and antibody-expressing lymphatic cells (eg, B cells) are recovered from the mice. Lymphoid cells can be fused with myeloma cell lines to produce immortal hybridoma cell lines, and such hybridoma cells to identify hybridoma cell lines that produce antibodies specific to the antigen of interest. Strains are screened and selected. The DNA encoding the heavy and light chain variable regions can be isolated and ligated to the desired isotypic constant regions of the heavy and light chains. Such antibody proteins can be produced in cells such as CHO cells. Alternatively, DNA encoding antigen-specific chimeric antibodies or light and heavy chain variable domains can be isolated directly from antigen-specific lymphocytes.

First, high affinity chimeric antibodies with human variable regions and mouse constant regions are isolated. Antibodies are characterized and selected for desired characteristics, including affinity, selectivity, epitope, etc. using standard procedures known to those of skill in the art. The murine constant regions are replaced with the desired human constant regions to generate the fully human antibodies featured in the invention, eg, wild-type or modified IgG1 or IgG4. Although the constant region selected may vary according to the particular application, high-affinity antigen-binding and target specificity characteristics reside in the variable region.

Generally, antibodies that may be used in the methods of the invention possess high affinities, as described above, as measured by binding to the antigen either immobilized on a solid phase or in solution phase. The mouse constant regions are replaced with the desired human constant regions to generate the fully human antibodies featured in this invention. Although the constant region chosen may vary according to the particular application, high-affinity antigen-binding and target specificity characteristics reside in the variable region.

Specific examples of antibodies or antigen-binding fragments of antibodies that specifically bind IL-4R that may be used in the context of the methods of the invention include SEQ ID NOs: 2, 18, 22, 26, 42, 46, 50, 66, 70, 74, 90, 94, 98, 114, 118, 122, 138, 142, 146, 162, 166, 170, 186, 190, 194, 210, 214, 218, 234, 238, 242, 258, and any antibody or antigen-binding fragment comprising the three heavy chain CDRs (HCDR1, HCDR2 and HCDR3) contained within the heavy chain variable region (HCVR) having an amino acid sequence selected from the group consisting of: The antibody or antigen-binding fragment has a , 178, 188, 192, 202, 212, 216, 226, 236, 240, 250, 260, and 264. Chain CDRs (LCVR1, LCVR2, LCVR3) may be included. Methods and techniques for identifying CDRs within HCVR and LCVR amino acid sequences are well known in the art and can be used to identify CDRs within the particular HCVR and/or LCVR amino acid sequences disclosed herein. obtain. Exemplary conventions used to identify CDR boundaries include, for example, the Kabat, Chothia, and AbM definitions. Broadly speaking, the Kabat definition is based on sequence variability, the Chothia definition is based on the location of structural loop regions, and the AbM definition is a compromise between the Kabat and Chothia approaches. See, eg, Kabat, "Sequences of Proteins of Immunological Interest," National Institutes of Health, Bethesda, Md. (1991); Al-Lazikani et al. , J. Mol. Biol. 273:927-948 (1997); and Martin et al. , Proc. Natl. Acad. Sci. See USA 86:9268-9272 (1989). Public databases are also available for identifying CDR sequences within antibodies.

In certain embodiments of the invention, the antibody or antigen-binding fragment thereof has , 70/72, 74/82, 90/92, 94/96, 98/106, 114/116, 118/120, 122/130, 138/140, 142/144, 146/154, 162/164, 166 /168, 170/178, 186/188, 190/192, 194/202, 210/212, 214/216, 218/226, 234/236, 238/240, 242/250, 258/260, and 262/ 6 CDRs (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3) from heavy and light chain variable region amino acid sequence pairs (HCVR/LCVR) selected from the group consisting of 264.

In certain embodiments of the invention, the antibody or antigen-binding fragment thereof is SEQ ID NO: 4/6/8/12/14/16; 28/30/32/36/38/40; 52/54/56/60 76/78/80/84/86/88; 100/102/104/108/110/112; 124/126/128/132/134/136; 148/150/152/156/158 172/174/176/180/182/184; 196/198/200/204/206/208; 220/222/224/228/230/232; and 244/246/248/252/254/ It contains 6 CDRs (HCDR1/HCDR2/HCDR3/LCDR1/LCDR2/LCDR3) having amino acid sequences selected from the group consisting of 256.

In certain embodiments of the invention, the antibody or antigen-binding fragment thereof has , 70/72, 74/82, 90/92, 94/96, 98/106, 114/116, 118/120, 122/130, 138/140, 142/144, 146/154, 162/164, 166 /168, 170/178, 186/188, 190/192, 194/202, 210/212, 214/216, 218/226, 234/236, 238/240, 242/250, 258/260, and 262/ HCVR/LCVR amino acid sequence pairs selected from the group consisting of 264.

Pharmaceutical Compositions The present invention includes methods comprising administering an IL-4R antagonist to a patient, wherein the IL-4R antagonist is contained within a pharmaceutical composition. The pharmaceutical compositions featured in the invention are formulated with suitable carriers, excipients, and other agents to provide proper transport, delivery, tolerance, and the like. Numerous suitable formulations can be found in formularies known to all pharmacists: Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, PA. These formulations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, lipid (cationic or anionic)-containing vesicles (e.g. LIPOFECTIN ^™ ), DNA conjugates, anhydrous absorbent pastes, oil-in-water and water-in-oil emulsions, emulsions carbowax (polyethylene glycols of various molecular weights), semi-solid gels, and semi-solid mixtures containing carbowax. Powell et al. See also "Compendium of excipients for parental formulations" PDA (1998) J Pharm Sci Technol 52:238-311.

The dose of antibody administered to a patient according to the methods of the present invention may vary according to the patient's age and size, symptoms, condition, route of administration, and the like. Preferred doses are typically calculated according to body weight or body surface area. Depending on the severity of the condition, the frequency and duration of treatment can be adjusted. Effective dosages and administration schedules for pharmaceutical compositions comprising anti-IL-4R antibodies can be determined empirically; for example, patient progress can be monitored by periodic assessment and dosages adjusted accordingly. Additionally, interspecies scaling of dosage may be performed using methods well known in the art (eg, Mordenti et al., 1991, Pharmaceut. Res. 8:1351).

Various delivery systems are known and can be used to administer the pharmaceutical compositions featured in the invention. For example, liposome encapsulation, microparticles, microcapsules, recombinant cells capable of expressing mutant viruses, receptor-mediated endocytosis (eg Wu et al., 1987, J. Biol. Chem. 262:4429- 4432). Methods of administration include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, intratracheal, epidural, and oral routes. The compositions may be administered by any conventional route, such as by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (eg, oral mucosa, rectal and intestinal mucosa, etc.), and others. of biologically active agents.

The pharmaceutical compositions of the invention can be delivered subcutaneously or intravenously using standard needles and syringes. Additionally, for subcutaneous delivery, pen-type delivery devices readily have utility in delivering the pharmaceutical compositions of the present invention. Such pen delivery devices may be reusable or disposable. Reusable pen delivery devices typically utilize replaceable cartridges containing pharmaceutical compositions. Once all of the pharmaceutical composition in the cartridge has been administered and the cartridge is emptied, the empty cartridge can be easily discarded and replaced with a new cartridge containing the pharmaceutical composition. The pen delivery device can then be reused. There are no replaceable cartridges in disposable pen delivery devices. Rather, the disposable pen device comes pre-filled with a pharmaceutical composition held in a reservoir within the device. When the reservoir is emptied of pharmaceutical composition, the entire device is discarded.

A number of reusable pen-type autoinjector delivery devices have utility in subcutaneous delivery of the pharmaceutical compositions of the present invention. Examples include, but are not limited to, AUTOPEN ^™ (Owen Mumford, Inc., Woodstock, UK), DISETRONIC ^™ pens (Disetronic Medical Systems, Bergdorf, Switzerland), HUMALOG MIX 75/25 ^{™ pens, HUMALOG™ pens} , to name a few. Pens, HUMALIN 70/30 ^™ pens (Eli Lilly and Co., Indianapolis, IN), NOVOPEN ^™ I, II and III (Novo Nordisk, Copenhagen, Denmark), NOVOPEN JUNIOR ^™ (Novo Nordisk, Copenhagen, Denmark), BD ^™ Pens (Becton Dickinson, Franklin Lakes, NJ), OPTIPEN ^™ , OPTIPEN PRO ^™ , OPTIPEN STARLET ^™ , and OPTICLIK ^™ (sanofi-aventis, Frankfurt, Germany). Non-limiting examples of disposable pen delivery devices that have utility in subcutaneous delivery of the pharmaceutical compositions of the present invention include the SOLOSTAR ^™ pen (sanofi-aventis), FLEXPEN ^™ (Novo Nordisk), and, to name a few. KWIKPEN ^™ (Eli Lilly), SURECLICK ^™ autoinjector (Amgen, Thousand Oaks, Calif.), PENLET ^™ (Haselmeier, Stuttgart, Germany), EPIPEN (Dey, LP), and HUMIRA ^™ pens (Abbott Labs, Abbott Park IL).

For direct administration to the paranasal sinuses, the pharmaceutical composition of the present invention can be administered through, for example, microcatheters (eg, endoscopes and microcatheters), aerosolizers.
r), may be administered using a powder dispenser, nebulizer or inhaler. The method includes administering an IL-4R antagonist to a subject in need thereof in an aerosolized formulation. For example, an aerosolized antibody to IL-4R can be administered to treat asthma in a patient. Aerosolized antibodies may be prepared, for example, as described in US8178098, which is incorporated herein in its entirety.

In certain circumstances, pharmaceutical compositions can be delivered in a controlled release system. In one embodiment, a pump may be used (see Langer, supra; Sefton, 1987, CRC Crit. Ref. Biomed. Eng. 14:201). In another embodiment, polymeric materials may be used; see, eg, Medical Applications of Controlled Release, Langer and Wise (eds.), 1974, CRC Press. , Boca Raton, Florida. In yet another embodiment, a controlled-release system can be placed in close proximity to the target of the composition, thus requiring only a fraction of the systemic dose (see, e.g., Goodson, 1984, in Medical Applications of Controlled Release, supra , vol.2, pp.115-138). Other controlled release systems are described in Langer, 1990, Science
249:1527-1533.

Injectable formulations may include dosage forms for intravenous, subcutaneous, intradermal and intramuscular injection, infusion, and the like. These injectable preparations can be manufactured by known methods. For example, injectable formulations can be prepared by dissolving, suspending or emulsifying the antibody or salt thereof in a sterile aqueous or oily medium conventionally used for injection. Aqueous vehicles for injection include, for example, saline, isotonic solutions containing glucose and other adjuvants, alcohols (eg, ethanol), polyalcohols (eg, propylene glycol, polyethylene glycol). , nonionic surfactants [eg Polysorbate 80, HCO-50 (polyoxyethylene (50 mol) adduct of hydrogenated castor oil)] and the like. Oily vehicles include, for example, sesame oil and soybean oil, which can be used in combination with solubilizers such as benzyl benzoate, benzyl alcohol, and the like. Injectables are therefore preferably filled into suitable ampoules.

Advantageously, the pharmaceutical compositions for oral or parenteral use described above are prepared into dosage forms in unit doses suitable to suit the dosage of the active ingredient. Such unit dose dosage forms include, for example, tablets, pills, capsules, injections (ampoules), suppositories, and the like.

Exemplary pharmaceutical compositions comprising anti-IL-4R antibodies that may be used in the context of the present invention are disclosed, eg, in US Patent Application Publication No. 2012/0097565.

Dosage The amount of IL-4R antagonist (eg, anti-IL-4R antibody) administered to a subject according to the methods of the invention is generally a therapeutically effective amount. As used herein, the phrase "therapeutically effective amount" means: (a) a reduction in the incidence of asthma exacerbations; (b) an improvement in one or more asthma-related parameters (defined elsewhere herein); and/or (c) an amount of IL-4R antagonist that produces one or more of detectable amelioration of one or more symptoms or signs of an upper airway inflammatory condition. A "therapeutically effective amount" also includes an amount of IL-4R antagonist that inhibits, prevents, reduces, or slows the progression of asthma in a subject.

For anti-IL-4R antibodies, the therapeutically effective amount is about 0.05 mg to about 600 mg of anti-IL-4R antibody, such as about 0.05 mg, about 0.1 mg, about 1.0 mg, about 1.5 mg, about 2.0 mg, about 3.0 mg, about 5.0 mg, about 7.0 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 110 mg, about 120 mg, about 130 mg, about 140 mg, about 150 mg, about 160 mg, about 170 mg, about 180 mg, about 190 mg, about 200 mg, about 210 mg, about 220 mg, about 230 mg, about 240 mg, about 250 mg, about 260 mg, about 270 mg, about 280 mg, about 290 mg, about 300 mg, about 310 mg, about 320 mg, about 330 mg, about 340 mg, about 350 mg, about 360 mg, about 370 mg, about 380 mg, about 390 mg, about 400 mg, about 410 mg, about 420 mg, about 430 mg, about 440 mg , about 450 mg, about 460 mg, about 470 mg, about 480 mg, about 490 mg, about 500 mg, about 510 mg, about 520 mg, about 530 mg, about 540 mg, about 550 mg, about 560 mg, about 570 mg, about 580 mg, about 590 mg, or about 600 mg could be. In a specific embodiment, 300 mg of anti-IL-4R antibody is administered.

The amount of IL-4R antagonist contained within an individual dose may be expressed in milligrams of antibody per kg of patient body weight (ie, mg/kg). For example, an IL-4R antagonist can be administered to a patient at a dose of about 0.0001 to about 10 mg/kg of patient body weight.

Combination Therapy According to certain embodiments, the methods of the invention comprise administering to the subject one or more additional therapeutic agents in combination with the IL-4R antagonist. As used herein, the phrase "in combination" means administering the additional therapeutic agent before, after, or concurrently with the pharmaceutical composition comprising the IL-4R antagonist. do. In some embodiments, the term "in combination" includes sequential or simultaneous administration of the IL-4R antagonist and the second therapeutic agent. The present invention includes a method for treating asthma or related conditions or complications, or for reducing at least one exacerbation, wherein the method is combined with a second therapeutic agent for additive or synergistic activity. administering an IL-4R antagonist.

For example, when administered "before" a pharmaceutical composition comprising an IL-4R antagonist, the additional therapeutic agent is administered about 72 hours, about 60 hours, about 48 hours before administration of the pharmaceutical composition comprising the IL-4R antagonist. hours before, about 36 hours before, about 24 hours before, about 12 hours before, about 10 hours before, about 8 hours before, about 6 hours before, about 4 hours before, about 2 hours before, about 1 hour before, about 30 hours before minutes before, about 15 minutes before, or about 10 minutes before. When administered “after” a pharmaceutical composition comprising an IL-4R antagonist, the additional therapeutic agent is administered 10 minutes, about 15 minutes, about 30 minutes after administration of the pharmaceutical composition comprising the IL-4R antagonist, After about 1 hour, after about 2 hours, after about 4 hours, after about 6 hours, after about 8 hours, after about 10 hours, after about 12 hours, after about 24 hours, after about 36 hours, after about 48 hours, It can be administered after about 60 hours, or after about 72 hours. Administration "simultaneously" with a pharmaceutical composition comprising an IL-4R antagonist means that the additional therapeutic agent is administered in a separate dosage form within less than 5 minutes (before, after, or at the same time) administration of the pharmaceutical composition comprising the IL-4R antagonist. or as a single combined dosage formulation containing both the additional therapeutic agent and the IL-4R antagonist.

Additional therapeutic agents include, for example, another IL-4R antagonist, IL-1 antagonists (including, for example, IL-1 antagonists described in US Pat. No. 6,927,044), IL-6 antagonists, IL-1 antagonists, IL-1 antagonists, 6R antagonists (including, for example, anti-IL-6R antibodies described in US Pat. No. 7,582,298), TNF antagonists, IL-8 antagonists, IL-9 antagonists, IL-17 antagonists, IL-5 antagonists , IgE antagonists, CD48 antagonists, leukotriene inhibitors, antifungals, NSAIDs, long-acting beta- ₂ agonists (e.g. salmeterol or formoterol), inhaled corticosteroids (e.g. fluticasone or budesonide), systemic corticosteroids ( for example, oral or intravenous), methylxanthine, nedocromil sodium, cromolyn sodium, or combinations thereof. For example, in certain embodiments, a pharmaceutical composition comprising an IL-4R antagonist is a combination comprising a long-acting beta ₂ agonist and an inhaled corticosteroid (eg, fluticasone + salmeterol [eg, Advair® ⁽ GlaxoSmithKline)) ]; or in combination with budesonide plus formoterol [eg Symbicort® ⁽ Astra Zeneca)]).

Dosing Regimens According to certain embodiments of the invention, multiple doses of the IL-4R antagonist may be administered to a subject over a defined time course. Such methods comprise sequentially administering multiple doses of the IL-4R antagonist to the subject. "Sequentially administering" as used herein means that each dose of the IL-4R antagonist is administered at different times, e.g., separated by a predetermined interval (e.g., hours, days, weeks, or months) , for example, means administered to a subject on different days. The present invention provides a single initial dose of an IL-4R antagonist, followed by one or more secondary doses of an IL-4R antagonist, and optionally followed by one or more tertiary doses of an IL-4R antagonist. and sequential administration to.

The present invention provides a pharmaceutical composition comprising an IL-4R antagonist to a subject about four times a week, twice a week, once a week, once every two weeks, for three weeks, as long as a therapeutic response is achieved. administering once every four weeks, once every five weeks, once every six weeks, once every eight weeks, once every twelve weeks, or less. In certain embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, once weekly dosing in amounts of about 75 mg, 150 mg, or 300 mg may be used. In other embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, biweekly dosing in amounts of about 75 mg, 150 mg, or 300 mg may be used. In other embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, once every three weeks dosing in amounts of about 75 mg, 150 mg, or 300 mg may be used. In other embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, a dose of about 75 mg, 150 mg, or 300 mg once every 4 weeks may be used. In other embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, dosages of about 75 mg, 150 mg, or 300 mg once every 5 weeks may be used. In other embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, once every 6 weeks dosing in amounts of about 75 mg, 150 mg, or 300 mg may be used. In other embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, once every 8 weeks dosing in amounts of about 75 mg, 150 mg, or 300 mg may be used. In other embodiments involving administration of a pharmaceutical composition comprising an anti-IL-4R antibody, dosages of about 75 mg, 150 mg, or 300 mg once every 12 weeks may be used. A preferred route of administration is subcutaneous.

The term "week" or "weeks" refers to a period of (n x 7 days) ± 2 days, preferably (n x 7 days) ± 1 day, more preferably (n x 7 days). where "n" designates the number of weeks, eg 1, 2, 3, 4, 5, 6, 8, 12 or more.

The terms "primary dose,""secondarydose," and "tertiary dose" refer to the timeline of administration of the IL-4R antagonist. Thus, the "initial dose" is the dose administered at the beginning of the treatment regimen (also called the "baseline dose"); the "secondary dose" is the dose administered after the initial dose; and the "tertiary dose." is the dose administered after the second dose. The initial, secondary, and tertiary doses may all contain the same amount of IL-4R antagonist, but may generally differ from one another in terms of frequency of administration. However, in certain embodiments, the amount of IL-4R antagonist contained in the initial, secondary and/or tertiary doses alternates (eg, is adjusted upwards or downwards as appropriate) over the course of treatment. In certain embodiments, two or more (e.g., 2, 3, 4, or 5) doses are administered as a "loading dose" at the beginning of the treatment regimen, followed by less frequent administration. Subsequent doses (eg, maintenance doses) are administered. In one embodiment, the maintenance dose can be lower than the loading dose. For example, one or more loading doses of 600 mg of IL-4R antagonist may be administered, followed by a maintenance dose of about 75 mg to about 300 mg.

In one exemplary embodiment of the invention, the second and/or tertiary dose, respectively, is 1 to 14 weeks (eg, 1, ¹ 1/2, ₂ , _{2 1/2} ^, 3 , 3 ^1/2 , ₄ , 4 ¹
/ ₂ , ⁵ , 5 ^1/2 , ₆ , 6 _1/2 , 7, 7 ^1/2 , 8, _{8 1/2} , 9, ^{9 1/2} , ₁₀ , ¹⁰ _1/2
, 11, 11 ^1/2 , _{12, 12 1/2 , 13, 13 1/2 , 14, 14 1/2 weeks} ^{or longer} ₎ ^. The phrase "previous dose" means a dose of an IL-4R antagonist administered to a patient in a series of multiple doses, without intervening doses, prior to administration of the next dose in that order.

These methods may comprise administering to the patient any number of secondary and/or tertiary doses of the IL-4R antagonist. For example, in certain embodiments, only a single secondary dose is administered to the patient. In other embodiments, two or more (eg, 2, 3, 4, 5, 6, 7, 8, or more) secondary doses are administered to the patient. Similarly, in certain embodiments, only a single tertiary dose is administered to the patient. In other embodiments, two or more (eg, 2, 3, 4, 5, 6, 7, 8, or more) tertiary doses are administered to the patient.

In embodiments involving multiple secondary doses, each secondary dose may be administered with the same frequency as the other secondary doses. For example, each secondary dose can be administered to the patient 1-2 weeks after the previous dose. Similarly, in embodiments involving multiple tertiary doses, each tertiary dose may be administered with the same frequency as the other tertiary doses. For example, each tertiary dose can be administered to the patient 2-4 weeks after the immediately preceding dose. Alternatively, the frequency with which the secondary and/or tertiary doses are administered to the patient may vary during the treatment regimen. Dosage frequency may also be adjusted during the course of treatment by a physician after clinical examination, depending on individual patient needs.

The present invention includes methods for treating asthma or related conditions comprising sequential administration of an IL-4R antagonist and a second therapeutic agent to a patient. In some embodiments, the invention provides one or more doses of an IL-4R antagonist followed by one or more doses of a second therapeutic agent (eg, 2, 3, 4, 5, 6, 7, 8, or more). For example, to treat, alleviate, decrease or ameliorate one or more symptoms of asthma, one or more doses of about 75 mg to about 300 mg of IL-4R antagonist may be administered, followed by a second One or more doses (eg, 2, 3, 4, 5) of a therapeutic agent (eg, an inhaled corticosteroid or beta 2-agonist or any other therapeutic agent described elsewhere herein) , 6, 7, 8, or more) may be administered. In some embodiments, the IL-4R antagonist is administered at one or more doses (e.g., 2, 3, 4, 5, 6, 7, 8, or more) followed by a second therapeutic agent to prevent recurrence of at least one asthma symptom. An alternative embodiment relates to co-administration of an IL-4R antagonist and a second therapeutic agent. For example, one or more doses (eg, 2, 3, 4, 5, 6, 7, 8, or more) of an IL-4R antagonist are administered, and the second therapeutic agent is I
It is administered in separate dosages with similar or different frequency as compared to the L-4R antagonist. In some embodiments, the second therapeutic agent is administered before, after, or concurrently with the IL-4R antagonist.

Treatment Population The methods of the invention involve administering a therapeutic composition comprising an IL-4R antagonist to a subject in need thereof. The phrase "a subject in need thereof" exhibits one or more symptoms or signs of asthma (e.g., eosinophilic asthma, including moderate to severe eosinophilic asthma) or has asthma means a human or non-human animal diagnosed with For example, a "subject in need thereof" may, for example, have a poor FEV1 (e.g., less than 2.0 L), a poor AM PEF (e.g., less than 400 L/min), a poor PM PEF (e.g., less than 400 L/min) prior to treatment. minutes), an ACQ5 score of at least 2.5, nocturnal awakenings at least once per night, and/or an SNOT-22 score of at least 20. (d) the subject. In various embodiments, these methods can be used to treat mild, moderate-to-severe and severe asthma in patients in need thereof.

In a related embodiment, the "subject in need thereof" has been treated with an inhaled corticosteroid (ICS)/long-acting beta2 _- adrenergic antagonist (LABA) prior to being administered an IL-4R antagonist. may have been prescribed or are currently taking a combination of Examples of ICS/LABA therapies include fluticasone/salmeterol combination therapy and budesonide/formotrol combination therapy. For example, the present invention provides for patients who have received the normal course of ICS/LABA for two weeks or more immediately prior to administration of the IL-4R antagonist (such prior treatment is referred to herein as "background treatment"). A method comprising administering an IL-4R antagonist to a patient who has had a disease. The present invention includes therapeutic methods in which the background treatment is discontinued at the time of the first administration of the IL-4R antagonist, or just prior to the first administration (eg, one day to two weeks prior). Alternatively, background treatment can be continued in conjunction with administration of an IL-4R antagonist. In still other embodiments, the amount of the ICS component, the amount of the LABA component, or both are gradually decreased before or after the initiation of IL-4R antagonist administration. In some embodiments, the invention includes methods for treating patients with persistent asthma for at least 12 months or longer. In one embodiment, patients with persistent asthma may be refractory to treatment with therapeutic agents such as corticosteroids and may be administered an IL-4R antagonist according to the invention.

In some embodiments, a "subject in need thereof" can be a subject with elevated levels of an asthma-related biomarker. Examples of asthma-related biomarkers include, but are not limited to, IgE, thymus and activation-regulated chemokines (TARC), eotaxin-3, CEA, YKL-40, and periostin. In some embodiments, a "subject in need thereof" can be a subject with a blood eosinophil level of 300/μl or greater or a sputum eosinophil level of 3% or greater. In one embodiment, a "subject in need thereof" can be a subject with elevated levels of bronchial or airway inflammation as measured by the fraction of exhaled nitric oxide (FeNO). .

For purposes of the present invention, normal IgE levels in healthy subjects are less than about 100 kU/L (e.g., ^ImmunoCAP® assay [Phadia, Inc. Portage,
MI]). Accordingly, the present invention provides for elevated serum IgE levels (greater than about 100 kU/L, greater than about 150 kU/L, greater than about 500 kU/L, greater than about 1000 kU/L, greater than about 1500 kU/L, about 2000 kU/L). higher than L,
Select subjects exhibiting serum IgE levels greater than about 2500 kU/L, greater than about 3000 kU/L, greater than about 3500 kU/L, greater than about 4000 kU/L, greater than about 4500 kU/L, or greater than about 5000 kU/L and administering to said subject a pharmaceutical composition comprising a therapeutically effective amount of an IL-4R antagonist.

TARC levels in healthy subjects range from 106 ng/L to 431 ng/L, with an average of about 239 ng/L. (An exemplary assay system for measuring TARC levels is the TARC Quantitative ELISA Kit provided by R&D Systems, Minneapolis, Minn., under catalog number DDN00.) Accordingly, the present invention provides a high, greater than about 500 ng/L, greater than about 1000 ng/L, greater than about 1500 ng/L, greater than about 2000 ng/L, greater than about 2500 ng/L, greater than about 3000 ng/L, greater than about 3500 ng/L , serum TARC levels greater than about 4000 ng/L, about 4500 ng/L, or greater than about 5000 ng/L, and a medicament comprising a therapeutically effective amount of an IL-4R antagonist It includes a method comprising administering the composition to a subject.

Eotaxin-3 belongs to a group of chemokines released into airway epithelial cells, which is upregulated by the Th2 cytokines IL-4 and IL-13 (Lilly et al 1999, J. Allergy Clin. Immunol. 104:786 -790). The present invention provides for patients with elevated eotaxin-3 levels, such as greater than about 100 pg/ml, greater than about 150 pg/ml, greater than about 200 pg/ml, greater than about 300 pg/ml, or greater than about 350 pg/ml. including administering an IL-4R antagonist to treat Serum eotaxin-3 levels are measured, for example, by ELISA.

Periostin is an extracellular matrix protein involved in Th2-mediated inflammatory processes. Periostin levels are found to be upregulated in patients with asthma (Jia et al 2012 J Allergy Clin Immunol. 130:647-654.e10.doi:10.1016/j.jaci.2012.06.025 .Epub 2012 Aug 1). The invention includes methods comprising administering an IL-4R antagonist to treat a patient with elevated levels of periostin.

Exhaled nitric oxide (Fractional exhaled NO) (FeNO) is a biomarker of bronchial or airway inflammation. FeNO is produced by airway epithelial cells in response to inflammatory cytokines, including IL-4 and IL-13 (Alwing et al.
1993, Eur. Respir. J. 6:1368-1370). FeNO levels range from 2 to 30 parts per virion (ppb) in healthy adults. An exemplary assay for measuring FeNO is by using a NIOX instrument by Aerocrine AB, Solna, Sweden. Assessments may be performed prior to spirometry and after fasting for at least 1 hour. The present invention provides IL in patients with elevated levels of exhaled NO (FeNO), such as greater than about 30 ppb, greater than about 31 ppb, greater than about 32 ppb, greater than about 33 ppb, greater than about 34 ppb, or greater than about 35 ppb. A method comprising administering a -4R antagonist is included.

Carcinoembryonic antigen (CEA) is a correlated tumor marker for lung non-neoplastic disease (Marechal et al 1988, Anticancer Res. 8:677-680). CEA levels in serum can be measured by ELISA. The present invention provides greater than about 1.0 ng/ml, greater than about 1.5 ng/ml, greater than about 2.0 ng/ml, greater than about 2.5 ng/ml, greater than about 3.0 ng/ml, about A method comprising administering an IL-4R antagonist to a patient with elevated levels of CEA, such as greater than 4.0 ng/ml, or greater than about 5.0 ng/ml.

YKL-40 [derived from its N-terminal amino acids tyrosine (Y), lysine (K) and leucine (L) and its molecular weight of 40 kD] is upregulated and correlated with asthma exacerbation, IgE, and eosinophils is a chitinase-like protein found in (Tang et al 2010 Eur. Respir. J. 35:757-760). Serum YKL-40 levels are measured, for example, by ELISA. The present invention provides for YKL-, such as greater than about 40 ng/ml, greater than about 50 ng/ml, greater than about 100 ng/ml, greater than about 150 ng/ml, greater than about 200 ng/ml, or greater than about 250 ng/ml. A method comprising administering an IL-4R antagonist to a patient with elevated levels of IL-40.

Induced sputum eosinophils and neutrophils are well-established direct markers of airway inflammation (Djukanovic et al 2002, Eur. Respire. J. 37:1S-2S). Sputum is induced with hypertonic saline inhalation and processed for cell counting according to methods known in the art, eg, European Respiratory Society guidelines. The invention includes methods comprising administering an IL-4R antagonist to a patient with elevated levels of sputum eosinophils, such as greater than about 2.5%, or greater than about 3%.

Methods for Evaluating Pharmacodynamic Asthma-Related Parameters Includes methods for assessing one or more pharmacodynamic asthma-related parameters. A reduction in the incidence of asthma exacerbations (as described above) or an improvement in one or more asthma-related parameters (as described above) may be correlated with an improvement in one or more pharmacodynamic asthma-related parameters. such correlations are not necessarily observed in all cases.

Examples of "pharmacodynamic asthma-related parameters" include, for example: (a) biomarker expression levels; (b) serum protein and RNA analysis; (c) induced sputum eosinophils and neutrophils. levels; (d) exhaled nitric oxide (FeNO); and (e) blood eosinophil counts. An "improved pharmacodynamic asthma-related parameter" is, for example, a decrease from baseline in one or more biomarkers such as TARC, eotaxin-3 or IgE, sputum eosinophils or neutrophils, FeNO , or decreased blood eosinophil count. As used herein with respect to pharmacodynamic asthma-related parameters, the term "baseline" refers to pharmacodynamic asthma-related parameters for a patient prior to or at the time of administration of a pharmaceutical composition featured in the invention. means the numerical value of

To assess pharmacodynamic asthma-related parameters, the parameters are determined at baseline and at time points after administration of the pharmaceutical composition of the invention. For example, pharmacodynamic asthma-related parameters were measured at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 days after initial treatment with a pharmaceutical composition of the invention. days, 11 days, 12 days, 14 days, or 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks 13 weeks, 14 weeks, 15 weeks, 16 weeks, 17 weeks, 18 weeks, 19 weeks, 20 weeks, 21 weeks, 22 weeks, 23 weeks, 24 weeks, or later can be measured. The difference between the value of the parameter at a specified time point after initiation of treatment and the value of the parameter at baseline is defined as an 'improvement'-like change in a pharmacodynamic asthma-related parameter (e.g., the specific parameter measured used to establish whether there was an increase or decrease as the case may be).

In certain embodiments, administration of an IL-4R antagonist to a patient causes a change, such as a decrease or an increase, in the expression of particular biomarkers. Asthma-related biomarkers include: (a) total IgE; (b) thymic and activation-regulated chemokines (TARC); (c) YKL-40; and (d) carcinoembryonic antigen (CEA) in serum. , also known as CEA cell adhesion molecule 5 [CEACAM5]), and (e) eotaxin-3 in plasma. For example, administration of an IL-4R antagonist to an asthmatic patient can cause one or more of a decrease in TARC or eotaxin-3 levels, or a decrease in total serum IgE levels. A decrease can be detected 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, or more after administration of the IL-4R antagonist. Biomarker expression can be assayed by methods known in the art. For example, protein levels can be measured by ELISA (enzyme-linked immunosorbent assay), or RNA levels by reverse transcription coupled with polymerase chain reaction (RT-PCR).

As discussed above, biomarker expression can be assayed by detection of protein or RNA in serum. Serum samples also monitor additional protein or RNA biomarkers associated with treatment with IL-4R antagonists, IL-4/IL-13 signaling, response to asthma, atopy or eosinophilic disease (e.g., soluble by measuring IL-4Rα, IL-4, IL-13, periostin). In some embodiments, RNA samples are used to determine RNA levels, e.g., biomarker RNA levels (non-genetic analysis); and in other embodiments, RNA samples are subjected to transcriptome sequencing (e.g., genetic analysis).

The following examples are presented to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the methods and compositions featured in the invention, and are intended to assist the inventors in their It is not intended to limit the scope of what is considered an invention of Efforts have been made to ensure accuracy with respect to numbers used (eg amounts, temperature, etc.) but some experimental errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, molecular weight is average molecular weight, temperature is in degrees Celsius, and pressure is at or near atmospheric.

Example 1. Generation of Human Antibodies Against Human IL-4R Human anti-hIL-4R antibodies were generated as described in US Pat. No. 7,608,693. Table 1 provides sequence identifiers for heavy and light chain variable region amino acid sequence pairs and CDR amino acid sequences of selected anti-IL-4R antibodies and their corresponding antibody designations.

An exemplary IL-4R antagonist used in the examples below is the human anti-IL-4R antibody designated H1H098-b in Table 1 (also referred to herein as "mAb1").

Example 2: Anti-IL-4R Antibody (mAb1) Administered Subcutaneously in Patients With Persistent Moderate to Severe Eosinophilic Asthma, Including Asthma Patients With Chronic Hypertrophic Eosinophilic Sinusitis A clinical trial of A. Study Objectives and Overview: A randomized, placebo-controlled, double-blind, parallel-group study was partially controlled/controlled with inhaled corticosteroid (ICS) and long-acting beta-2 agonist (LABA) treatment. Patients with persistent moderate-to-severe eosinophilic asthma who did not receive either 300 mg mAb1 or placebo subcutaneously once weekly for 12 weeks. The primary objective of the study was the reduction in the incidence of asthma exacerbations in patients with persistent moderate-to-severe eosinophilic asthma of mAb1 administered subcutaneously once weekly for 12 weeks compared to placebo. The purpose was to investigate the effect. The secondary objectives of this study were to assess the safety and tolerability of mAb1 administered subcutaneously once weekly for 12 weeks in patients with persistent moderate-to-severe eosinophilic asthma and to evaluate mAb1 serum concentrations after weekly subcutaneous dosing for 12 weeks in patients with moderate to severe eosinophilic asthma.

Prior to Screening, patients were required to have been on a stable dose of any of the following doses and regimens of ICS/LABA combination therapy (also referred to as "background therapy") for at least 1 month:
Fluticasone/Salmeterol Combination Therapy - Advair ^® Diskus - Dry Powder Inhaler (DPI): 250/5
0 ug twice daily (BID) or 500/50 ug BID; or - Advair HFA - Metered dose inhaler (MDI ⁾ : 230/42 ug BID or 460/42 ug BID; or Budesonide/formoterol combination therapy (Symbicort ^{( R)} 160/9u
g
BID or 320/9ug BID); or mometasone/formoterol combination therapy ( ^Dulera® 200/10ug B
ID or 400/10ug BID).

Patients on budesonide/formoterol or mometasone/formoterol were switched to an equivalent dose of fluticasone/salmeterol at randomization (Day 1) and patients on fluticasone/salmeterol were on background treatment remained the same.

Patients who met the inclusion and exclusion criteria (see below) were randomized to one of the following treatments: mAb1 300 mg subcutaneously once weekly for 12 weeks; or placebo for 12 weeks; It was administered subcutaneously once a week.

The study consisted of a 2-week screening period, a 12-week treatment period including a 4-week background treatment stabilization period and an 8-week background treatment washout period after randomization, followed by an 8-week post-treatment follow-up period. included.

Algorithm for background treatment (ICS/LABA) withdrawal:
Patients remained on twice daily (BID) fluticasone/salmeterol background therapy for 4 weeks after starting adjunctive therapy with 300 mg mAb1 (or placebo) or treatment. After 4 weeks of randomization, patients were transferred from BID fluticasone/salmeterol combination therapy to equivalent ICS doses of fluticasone monotherapy (250ug or 500ug Flovent ^(R) Diskus-BIDDPI formulation; or 220ug or 440ug BID Flovent ^(R) HFA). - including any of the MDI formulations). The LABA component (ie salmeterol) was discontinued. If at subsequent visits the patient did not meet any of the criteria for asthma exacerbation (as defined below), the fluticasone dose was reduced by approximately 50% beginning at week 6. If asthma exacerbation was desired, ICS withdrawal proceeded according to the following schedule:

Upon completion of 12 weeks of treatment with study drug (or after early discontinuation), patients were placed on fluticasone/salmeterol to control symptoms for an additional 8 weeks of off-study medication prior to final safety assessment. , budesonide/formoterol, or mometasone/formoterol (doses at study entry) and their original doses of albuterol or lev-albuterol as appropriate.

Adult patients were included in the study based on the following criteria: (1) Physician's diagnosis of persistent asthma for at least 12 months based on the Global Initiative for Asthma (GINA) 2009 guidelines; , the airway inflammation may be eosinophilic; and (2) whether the asthma is partially controlled with inhaled corticosteroid/long-acting beta-agonist combination therapy according to the following criteria: or not controlled: (i) fluticasone/salmeterol combination therapy (DPI formulation: 250/50 μg BID or 500/50 μg BID or MDI: 230/42 μg BID or 460/42 μg BID) for at least 1 month prior to screening, or budesonide/ stable dose of either formoterol combination therapy (160/9 μg BID or 320/9 μg BID) or mometasone/formoterol combination therapy (200/10 μg BID or 400/10 μg BID); (iii) a Juniper Asthma Control Questionnaire (5-question version, ACQ) score of ≥1.5 and ≤3.0 at screening; Predicted normal with FEV1 ≥ 50% during the screening phase (up to 3 trials) and at randomization prior to first dose (up to 3 trials); (v) within 2 years prior to screening either treatment with one or more systemic (oral and/or parenteral) steroid bursts for asthma exacerbation, or hospitalization or emergency care visit for asthma exacerbation; and (vi) screening. Documented reversibility within 12 months of criteria-at least 12% and 200 mL (up to 3 trials) in FEV1 after albuterol 200 μg-400 μg (2-4 inhalations) during the screening phase History, or documented history of positive methacholine load (PD20 methacholine <8 mg) within 12 months prior to screening. Inhaled corticosteroids and long-acting beta agonists ⁽ ADVAIR®, SYMBIC
ORT ^(R) or DULERA ^(R) ) partially controlled or uncontrolled with moderate to high dose combination therapy and equal to or greater than 300 cells per microliter during the screening phase. Patients with blood eosinophils greater than or equal to 3% or sputum eosinophils equal to or greater than 3% were included in the study.

Patients meeting all inclusion criteria were screened for the following exclusion criteria: (1) patients <18 years or >65 years; (2) clinical findings suggestive of unknown disease and requiring further evaluation; (3) chronic obstructive pulmonary disease (COPD) and/or other pulmonary diseases that impair pulmonary function tests; (4) beta-adrenergic receptor blockers for any reason; (5) Current smoker or quit within 6 months prior to Screening; (6) History of smoking >10 packs per year; (7) Hospitalization or hospitalization due to asthma exacerbation within 2 months prior to Screening; (8) plans to initiate allergen immunotherapy within the study period; (9) a pre-screening period of less than 5 antibody half-lives but ≥30 days or known antibody half-life; If not, exposure to another study antibody within the pre-screening period that is at least 6 months; (10) previous enrollment in the current study; (12) known or suspected non-compliance, alcohol or substance abuse; (13) inability to follow test procedures (e.g., language problems or psychological (14) reversal of sleep patterns (e.g. night workers); (15) treatment with drugs known to prolong the QTc interval; (16) ICS (e.g. (17) within 2 months prior to screening, or use of more than 3 courses of injectable or oral systemic glucocorticoids within 6 months prior to screening; (18) alone or in combination with a nonsteroidal controller (fluticasone/salmeterol combination therapy, (19) patients receiving prohibited concomitant medications (listed below); (20) doxycycline or related (21) unwillingness to become pregnant, breast-feed, or use effective methods of contraception during pregnancy or the study; and (22) recent history of parasitic infection or 6 months prior to screening. Travel to parasite-endemic areas within .

Patients remained on a constant dose of background asthma therapy for the first 4 weeks of the study, then
Gradually decrease the dose of background therapy. Initially, the long-acting beta-agonist component of background therapy was withdrawn at week 4, then the inhaled corticosteroid dose was halved every two weeks for up to 12 weeks. Patients continued on study treatment until the end of the study or until withdrawal due to asthma exacerbation or any other reason.

B. Study Treatment Study Drug: Sterile mAb1 150 mg/mL solution for subcutaneous (SC) injection was provided in 5 mL glass vials. Each vial contained 2 mL of withdrawable volume. A 300 mg dose was administered subcutaneously at the test site once weekly in the morning for 12 weeks.
Placebo: A sterile placebo for SC injection was provided in a perfectly matched 5 mL glass vial. Each vial contained 2 mL of drawable volume. Placebo was administered subcutaneously at the test site once weekly in the morning for 12 weeks.

The following concomitant medications were not allowed during the study period: fluticasone/salmeterol combination therapy or fluticasone administered per the protocol (or budesonide/formoterol or mometasone/ any other inhaled steroid other than formoterol); systemic or ocular steroids; LABA other than the salmeterol component of fluticasone/salmeterol combination therapy administered according to the protocol; any other ICS/ any inhaled anticholinergic (e.g. ipratropium bromide or tiotropium); methylxanthines (theophylline, aminophylline); cromones; anti-IgE therapy; lipoxygenase inhibitors; agent.

C. Efficacy of Treatment The primary endpoint of this study was the development of asthma exacerbations, defined by either: (1) improvement in morning peak expiratory flow (PEF) from baseline on 2 consecutive days; or (2) 6 or more additional seizure drug puffs of albuterol or lev-albuterol (compared to baseline) in 24 hours on 2 consecutive days; or (3) ( Trial that required a) systemic (oral and/or parenteral) steroid treatment, or (b) a 4-fold or greater increase in the last dose of ICS received prior to discontinuation from the study, or (c) hospitalization Asthma exacerbation as determined by the attending physician.

Secondary endpoints of the study included the mean change from baseline in the following parameters: assessed at baseline and at treatment endpoint (week 12) to assess upper respiratory tract symptoms, (1 (2) morning and evening peak expiratory flow (AM PEF and PM PEF) in liters/min measured daily; (3) (4) five-item Asthma Control Questionnaire (ACQ5) score by visit; and (5) nocturnal awakenings measured daily (per night). and (6) a 22-item sinus assessment test (SNOT-22). Secondary endpoints also included 6 or more additional seizure medication puffs of albuterol or lev-albuterol (compared to baseline) in 24 hours on 2 consecutive days, and baseline in PEF in the morning on 2 consecutive days. The proportion of patients with combined asthma events defined by a 30% or greater reduction from line was included. PEF, ACQ5, Asthma Symptom Score, Nocturnal Arousal, and Seizure Medication Treatment Use were obtained by electronic diary. Mean daily nocturnal awakenings ranging from 0 to 10 were averaged from the previous 7 days. Morning and evening asthma symptom scores consisted of unvalidated patient-reported outcomes assessed on a 5-point Likert-type scale, with higher scores indicating worse outcomes (Table 2). ). Patients recorded global symptom scores twice daily before measuring PEF. Data are presented as the mean of the 7 days prior to the specified time point (e.g., Figure 26
A and 26B).

D. Adverse Event Monitoring Safety was assessed throughout the study by monitoring adverse events and serious adverse events.

An Adverse Event (AE) is any adverse medical occurrence in a subject administered a drug or in a clinical study subject. Therefore, an AE is any unfavorable unintended sign (abnormal laboratory ), symptom, or disease. AEs may also be: any worsening of a pre-existing condition (i.e., any clinically significant change in frequency and/or intensity) temporally related to study drug use; and any adverse medical event.

A Serious Adverse Event (SAE) results in death at any dose; is life-threatening; requires hospitalization or extension of current hospitalization; persistent or significant disability. is a congenital anomaly/defect; or is a serious medical event.

E. Statistical Methods For the primary analysis of the proportion of patients experiencing asthma exacerbations, logistic regression models were used to compare the SAR group to placebo. The model included terms for treatment and stratification factors (preceding ICS/LABA combination therapy dose). The primary analysis was based on the modified intent-to-treat (mITT) population, which included all randomized patients who received at least one dose of the investigational drug (IMP). I was there. A stratified chi-square test was also used to confirm the primary analysis.

For secondary efficacy endpoints, except for SNOT-22, changes from baseline were analyzed using a mixed effects model with a repeated measures (MMRM) approach. This model includes changes from baseline values through week 12 as response variables and the interaction of factors for treatment (parameter effect), stratification factors, visits, treatment-by-visit. , baseline values, and baseline-by-visit interactions. Statistical estimates for treatment comparisons for change from baseline at Week 12 were derived from a mixed effects model. Changes from baseline in SNOT-22 were analyzed using analysis using analysis of covariance (ANCOVA), with end-of-treatment measurements to impute missing data. Pharmacodynamic effects were assessed using the MMRM model in a post-hoc manner. No adjustment for multiplicity was made as there was only one primary endpoint and analysis. Safety variables including AEs, laboratory parameters, vital signs, ECG, clinical laboratory findings, and physical examination were summarized using descriptive statistics.

Demographic and clinical characteristics were summarized using descriptive characteristics. Plots of secondary and pharmacodynamic variables are shown as mean change from baseline over time with standard errors. Comparison of treatment effects from MMRM analysis is based on least squares mean change from baseline at week 12 (95% confidence interval [CI]).

F. Results The results observed in all 104 randomized patients (491 screened) who completed or discontinued the treatment period of the study are summarized below. All randomized patients were exposed to study treatment and included in the mITT population. Baseline characteristics were similar between groups. Demographic and clinical characteristics were also similar between the two groups (Table 3). Patients were treated with either weekly 300 mg subcutaneous mAb1 or placebo as indicated above. The study treatment period was completed by 86.5% and 67.3% of mAb1 and placebo patients, respectively (Figure 25). The most common cause of discontinuation was lack of efficacy, which was more frequent with placebo (21.2%) than with mAb1 (1.9%).

(i) Primary Efficacy Endpoint The incidence of asthma exacerbations in the placebo and mAb1 treated groups is shown in Table 4.

There were a total of 26 asthma exacerbations during the treatment period and no patients were hospitalized for asthma exacerbations. Twenty-three patients (44.2%) experienced asthma exacerbations in the placebo group, while only three patients (5.8%) experienced asthma exacerbations in the mAb1-treated group. The odds ratio was 0.077 (p<0.0001) and the relative risk reduction was approximately 87%.

The lack of this study, as demonstrated by the need for immediate intervention in the form of treatment with either systemic corticosteroids or inhaled corticosteroids at doses four times or greater than the dose ingested prior to the event. Of the 26 asthma exacerbations experienced during the period, 9 were considered severe. A summary of the incidence of severe asthma exacerbations is shown in Table 5.

As shown in Table 5, 8 severe asthma exacerbations were seen in the placebo group and only 1 severe asthma exacerbation in the mAb1 treated group. The remaining 15 asthma exacerbations in the placebo group and 2 in the mAb1 group meet the protocol definition of an exacerbation based on decreased morning PEF and/or increased albuterol/lev-albuterol use. As shown in Table 6, within the active treatment group, sustained improvements over baseline were observed during the course of the study for all parameters despite steroid withdrawal.

With mAb1, the time to exacerbation was longer compared to placebo (Fig. 1) and the risk of exacerbation was reduced (hazard ratio 0,10; 95% CI 0.03, 0.34; P<0. 001). Analysis of time to asthma exacerbation by Kaplan-Meier plot shows that the effect of treatment with mAb1 persists for longer periods, including 8 weeks after patients are at higher risk of exacerbations due to steroid withdrawal. (Fig. 1).

Only one patient from the placebo group had a combined asthma event. A composite asthma event was defined as a reduction from baseline in the morning PEF on 2 consecutive days with 6 or more additional seizure medication puffs of albuterol or levalbuterol (compared to baseline) in 24 hours on 2 consecutive days. Defined as a reduction of 30% or greater.

(ii) Other Efficacy Endpoints Pulmonary function parameters (FEV1, AM PEF and PM PEF), asthma symptom-based endpoints (ACQ score, nocturnal awakening), and albuterol use were assessed for each patient at each visit. . Observed results (weekly changes from baseline) for these parameters are shown in Figures 2-7, respectively. In addition, SNOT-22 scores were assessed at baseline and end of treatment. For all parameters, baseline and Week 12 (LOCF) mean values are summarized in Table 7, along with mean differences between treatment groups (ANOVA model for SNOT-22). In Table 7, the column labeled "Difference vs. Placebo" is the placebo correction from baseline to account for the observed change in the value of the parameter compared to the observed change for that parameter in the placebo-treated group. indicate a value.

Treatment with mAb1 resulted in a significant change from baseline in FEV1 at week 1, which was maintained through week 12 despite LABA and ICS withdrawal (Fig. 2), and 5 There was a slight decrease in FEV1 at week 1. A similar improvement was observed in the morning PEF, but less so in the evening PEF (Figures 3 and 4). The least squares (LS) mean change from baseline to week 12 in FEV1 was −0.22 L for placebo and 0.05 L for the mAb1 group (p=0.0009).

ACQ5 scores improved in both treatment groups at week 1 (Figure 6). However, although ACQ5 improved further with mAb1 between weeks 1 and 4, the placebo effect stabilized and maintained a difference up to 12 weeks.

Morning symptom scores increased from baseline to week 12 with placebo. With mAb1, there was an initial decline that remained below baseline by week 12 (FIG. 26A). A similar pattern (greater variability) was observed for nighttime asthma scores (Fig. 26B).

Nocturnal awakening was stable from the placebo group until week 6, then increased from week 6 to week 12. In contrast, nocturnal awakening decreased in the mAb1 group by week 1 and remained improved relative to baseline by week 12 (FIG. 7).

Change in albuterol/lev-albuterol use (Figure 5) was similar to other secondary endpoints: return to baseline after initial decline with placebo. For mAb1, the initial decline was maintained over time.

There was a non-significant difference at baseline between SNOT-22 values, with a mean placebo score of 26.24 and a mean mAb1 score of 39.02. At week 12, the LS mean change was a slight increase of 0.23 points for the placebo group and a mean decrease (improvement) of 8.26 points for the mAb1 group. This corresponds to an improvement magnitude of 8.49 points for the mAb1 group (p=0.0027).

For all secondary endpoints, with the exception of nocturnal PEF and nocturnal wakefulness, week 12 measurements were favorable and significant with mAb1 treatment (Tables 7 and 8). A significant improvement with mAb1 was also observed for three SNOT-22 items associated with upper respiratory tract disease (Table 9).

(iii) Safety mAb1 is generally safe and well tolerated. Treatment-emergent adverse events (TEAEs) were similarly reported by 40 (76.9%) placebo-treated and 42 (80.8%) mAb1-treated patients (Table 10). TEAEs were nonspecific, generally mild to moderate in intensity, and mostly resolved by the end of the study. The following increased reports of TEAEs were seen for mAb1 compared to placebo: Injection site reactions were reported by 15 (28.8%) mAb1 patients and 5 (9.6%) placebo patients nasopharyngitis was reported by 7 (13.5%) mAb1 patients and 2 (3.8%) placebo patients; headache was reported by 6 (11.5%) mAb1 patients and 3 (3.8%) 5.85) were reported by placebo patients, and nausea was reported by 4 (7.7%) mAb1 patients and 1 (1.9%) placebo patient.

No deaths were reported during the study period. Four treatment-emergent serious adverse events (SAEs) reported: 1 mAb1 patient experienced bipolar disorder and 3 placebo patients had asthma with pneumonia, gunshot wound with left pneumothorax , and an SAE of a right ankle fracture. None of these SAEs were considered related to IMP, and all but new ankle fractures resolved by the end of the study. There were no deaths.

A total of 6 patients discontinued the study due to TEAEs: 3 patients in the mAb1 group (bipolar disorder, asthma with wheezing, and angioedema) and 3 patients in the placebo group (upper respiratory tract infection, psoriasis and asthma). Angioedema TEAE occurred in a 42-year-old African American female after the ninth dose of study treatment as a pruritic general rash observed at and away from the injection site. This persisted for 1 week and resolved after study treatment discontinuation and prednisone and diphenhydramine treatment. This was considered treatment related. This AE was after a milder rash at the injection site after the first and sixth study treatment doses.

Among the most common AEs occurring in 3 or more patients in either treatment group (Table 10), injection site reactions, nasopharyngitis, nausea and headache occurred more frequently with mAb1 than with placebo. Clinically significant changes in vital signs, physical examination, laboratory tests or ECG findings are reported in either group.

G. Conclusions Significant improvements in pulmonary function and other asthma management parameters were observed. Efficacy was observed early and persistently despite background treatment withdrawal. An approximately 87% relative reduction (p<0.0001) in the primary endpoint of the incidence of asthma exacerbations in patients with persistent moderate-to-severe asthma with eosinophilia compared with placebo (44.2%) ) after 12 weeks of treatment with mAb1 300 mg once weekly (5.8%). As shown in Table 7, clinically meaningful and statistically significant (no multiplicity adjustment) improvements with treatment compared to placebo were observed in pulmonary function parameters (FEV1, PEF AM), asthma symptom score (ACQ ) and albuterol use. A beneficial trend was observed for PEF PM (p=0.0567) and nocturnal awakening (p=0.0518). A statistically significant (unmultiplicity-adjusted) improvement was also observed for SNOT-22 scores. Within the active treatment group, sustained improvement over baseline was observed during the course of the study for all parameters despite withdrawal of LABA and ICS. mAb1 is generally safe and well tolerated.

Example 3: Biomarker Studies Biomarker analyzes were performed on samples taken from subjects who participated in clinical trials of mAb1 (see Example 2 above). Specifically, thymus and activation chemokine (TARC; CCL17), immunoglobulin E (IgE), eotaxin-3, periostin, carcinoembryonic antigen (CEA), YKL-40 and blood eosinophil Serum/plasma biomarkers associated with TH2 inflammation, such as spheres, were measured in patient-derived samples at baseline and at various time points after initiation of study treatment. Baseline levels of these biomarkers were evaluated for their potential predictive value for treatment response. In addition, exhaled NO (FeNO) and induced sputum eosinophils and neutrophils were measured as biomarkers of bronchitis. Exhaled nitric oxide assessment was performed using a NIOX instrument (Aerocrine AB, Solna, Sweden) prior to spirometry and after at least 1 hour of fasting. Biomarkers were analyzed using a mixed model and the least squares means derived from that model are reported below.

Asthmatic subjects (N=104) received either mAb1 (300 mg) or placebo on study days 1, 8, 15, 22, 29, 36, 43, 50, 57, 64, 71, and 78 ( ie, 12 weekly doses) were administered subcutaneously (see Example 2 herein). Samples for biomarker analysis were collected from antibody- and placebo-treated subjects at 0, 1, 4, 8 and 12 weeks. Antigen-specific IgE was detected using the ^Phdiatop® test.

TARC, eotaxin-3, and IgE remained unchanged in response to placebo (Figures 8, 9 and 10). In contrast, TARC (−22.7% versus −0.3% mean % change; p=0.0003) (FIG. 8) and Eotaxin-3 (−39. A rapid reduction of 62%; p<0.0001) (FIG. 9) was observed in patients treated with mAb1 within 1 week and persisted up to 12 weeks: TARC: +7.6% vs. placebo -26.0% (p=0.0005); Eotaxin-3: +5.13% -45.67% vs placebo (p<0.0001).

TARC levels responded within 1 week after exposure to mAb1 administered subcutaneously at 300 mg. TARC levels stabilized at approximately 50% of baseline levels in mAb1-treated subjects despite ICS withdrawal. This data suggests that TARC expression is more directly related to IL-4R signaling than FEV1 changes, which decreased in parallel with ICS withdrawal [after 4 weeks], and that IL-4R blockade induces a shift towards a TH1 signature, for example, as observed with IFN-gamma administration. It may be possible to titrate mAb1 doses using TARC (and eg CXCL10), especially in patients requiring long-term treatment and at risk for TH1-type immune disorders.

Total serum IgE also decreased after mAb1 treatment. Total serum IgE responses were more heterogeneous and delayed compared to TARC responses. Mean (SD) baseline IgE levels were 694.68 IU/L (1837.82) for the placebo group (n=52) and 657.66 (1482.25) for the mAb1 group (n=52), although , while the median was 169.95 for the placebo group and 206.15 for the mAb1 group. Despite this heterogeneity, a trend toward IgE reduction in mAb1-exposed patients compared to placebo was observed, beginning only at 4 weeks. Serum IgE was significantly reduced in the mAb1 group compared to placebo (mean % change, +10.1% vs. +13.5%; p=0.0325) starting at week 4 and continuing for 12 weeks. (mean % change, −36.8% for REGN668/SAR231893 vs −5.5% for placebo; p<0.0001) (FIG. 10).

Changes from baseline and placebo at week 12 for FeNO, TARC, eotaxin-3, and IgE all favored mAb1 (all P<0.001) (Table 11). No difference from baseline or between treatments was observed with YKL-40 or CEA.

There was a transient decrease in periostin levels followed by an increase with LABA/ICS withdrawal (Figure 11). Administration of mAb1 slowed the increase but did not prevent the increase over baseline. No consistent treatment effect was observed with CEA (Figure 12) and YKL-40 (Figure 13). The number of blood eosinophils remained unchanged until 6 weeks, but then increased at 8 and 12 weeks (Fig. 14). Peripheral blood eosinophil counts did not change with placebo throughout treatment. Differences between treatments were not significant, and borderline increases were facilitated by greater blood eosinophilia in only a few patients treated with mAb1. Little or no increase was observed in most of the patients (Table 12).

No conclusions could be drawn regarding the association between baseline biomarker levels and asthma exacerbations, as only 3 mAb1 patients experienced asthma exacerbations during the study.

mAb1 treatment was also associated with a significant decrease from baseline in FeNO at week 4, and despite ICS withdrawal, FeNo remained below baseline through week 12 (mean % change: −28.7 for mAb1 vs. 35.0 for placebo; p<0.0001) (FIG. 15). In contrast, placebo FeNo values remained stable through week 8 and then increased consistent with ICS withdrawal.

Improvement in forced expiratory volume in 1 second (FEV ₁ ) was associated with a decrease in FeNO at week 12 (r=−0.40
8, p=0.009) (Fig. 16). Similarly, improvements in AM-PEF and PM-PEF correlated with FeNO reduction (FIGS. 17 and 18). Other correlations with FeNO were not significant. See Table 13.

Scatterplot analysis of change from baseline in Week 12 baseline eosinophils vs. FEV1 showed baseline eosinophils versus treatment effect, as measured by change from baseline in FEV1 at Week 12 in the study population. (baseline eosinophils > 0.3 giga/L) (Figure 19). Baseline eosinophils correlated with decreased ACQ (Figure 20) and decreased albuterol/levalbuterol use (Figure 21). Periostin and YKL-40 at baseline correlated with decreased ACQ (Figures 22 and 23).

FEV1 change from baseline at Week 12 was exacerbated by ICS withdrawal (beginning at Week 4). A similar analysis showed no association between baseline TARC or IgE and change from baseline in FEV1 at week 12 in the study population (baseline eosinophils > 0.3 giga/L) .

Summary These results show that mAb1 significantly reduced serum biomarkers associated with Th2 inflammation (TARC, eotaxin-3 and IgE) and bronchial inflammation (FeNO) in adult asthmatics. A correlation between FeNO reduction and FEV ₁ improvement was
suggest a relationship between IL-13-mediated anti-inflammatory activity and improved lung function in moderate to severe uncontrolled asthma.

The present invention should not be limited in scope by the specific embodiments described herein. Indeed, various modifications in addition to those described herein will become apparent to those skilled in the art from the foregoing description and accompanying drawings. Such modifications are intended to fall within the scope of the appended claims.

Example 4 Blockade of the IL-4/IL-13 Signaling Pathway Inhibits IgE Production and Airway Remodeling in a Mouse Model of House Dust Mite-Induced Eosinophilic Asthma and IL-4-induced transendothelial migration of eosinophils into the lung to induce Th2 immune responses. Eosinophils are the major effector cells in allergic reactions, and release of granular components (including IL-4) from eosinophils contributes to inflammation. In patients with asthma, Th2-driven production of IL-4 promotes blood-to-lung eosinophil migration via eotaxin, a potent eosinophil chemoattractant (Mochizuki et al., J. Immunol., 1998, 160(1):60-68). Furthermore, when localized to sites of inflammation, eosinophils produce and secrete IL-4, thus contributing to ongoing Th2-driven inflammation (Bjerke et al., Respir. Med., 1996, 90 (5):271-277). In patients with allergic asthma, HDM challenge increased levels of serum IgE and Th2 cytokines by 5 weeks after allergen challenge (van de Pol et al., Allergy, 2012, 67(1):67 -73).

In this example, the HDM-induced model of chronic asthma was used to assess the pharmacodynamic effects of anti-IL-4R antibodies on markers of airway inflammation in mice. Furthermore, since collagen deposition correlates with the degree of airway remodeling, the effect of anti-IL-4R antibodies on collagen deposition in the airways was evaluated in this model.

Materials and Methods Two different anti-IL-4Rα antibodies were used in the experiments of this example: “mAb1,” a fully human monoclonal antibody specific for human IL-4Rα (i.e. anti-IL-4R antibody used in the examples); and "anti-mIL-4Rα," a mouse monoclonal antibody specific for the mouse IL-4Rα protein. mAb1 does not cross-react with murine IL-4Rα; therefore, mAb1 is a humanized mouse (IL-4Rα) engineered to replace both the ectodomains of human IL-4 and IL-4Rα with the corresponding mouse sequences in the mouse (IL-4Rα). 4 ^hu/hu IL-4Rα ^hu/hu ). On the other hand, the mouse anti-mouse IL-4Rα antibody "anti-mIL-4Rα" was tested in wild-type (Balb/c) mice. A murine IL-13Rα2-mFc fusion protein acting as a decoy receptor that blocks IL-13 signaling by sequestration of IL-13 cytokines was also tested in these experiments.

For the HDM-induced asthma model, mice were sensitized with a daily intranasal application of HDM (50 μg in 20 μL PBS per mouse) for 10 days followed by rest (2 weeks recovery period). Allergen challenge was administered by intranasal application of HDM (50 μg in 20 μL PBS per mouse) three times weekly for 8 weeks. For each dose of HDM, mice were lightly anesthetized with isoflurane during either the sensitization or exposure period.

Mice were acclimated in the experimental facility for a minimum of 5 days prior to initiation of experimental procedures. During the entire duration of the experiment, animals remained housed in the experimental facility with ad libitum access to food and water on a 12-hour light/dark cycle under standard conditions. The number of mice per cage was limited to a maximum of 5 mice.

A total of 48 humanized mice (IL-4 ^hu/hu IL-4Rα ^hu/hu ) engineered to replace the human IL-4 ligand and the human ectodomain of IL-4Rα with their corresponding mouse sequences were subdivided into two groups. Used for experiments. IL-4 ^hu/hu IL-4Rα ^hu/hu mice are on a mixed background C57Bl/6NTac (75%)/129S6SvEvTac (25%). Additionally, 20 wild-type littermate mice of the same mixed background were used in one of three experiments. In each experiment, mice were sensitized with HDM (or PBS in the control group) daily for 10 days, followed by rest from day 11 to day 29. From day 30, animals were exposed to HDM three times a week for 8 weeks until day 81, then euthanized for analysis on day 85. Mice were divided into 6 experimental groups as follows:
(1) Non-sensitized, untreated : PBS was applied intranasally during the sensitization and exposure period. Mice were not treated with antibodies (IL-4 ^hu/hu IL-4Rα ^hu/hu mice, n=9; wild-type littermates n=5);
(2) HDM sensitization, no treatment : HDM was applied intranasally during the sensitization and exposure period. Mice were not treated with antibodies (IL-4 ^hu/hu IL-4Rα ^hu/hu mice, n=7; wild-type littermates, n=5);
(3) HDM sensitization, treatment with anti-mIL-4Rα : HDM was applied intranasally during the sensitization and exposure period. Mice were injected intraperitoneally (ip) with anti-mIL-4Rα at a dose of 50 mg/kg twice weekly from 7 weeks to 12 weeks for a total of 12 doses for a period of 6 weeks (wild-type littermates, n=5);
(4) HDM sensitization, treatment with anti-human mAb1 : HDM was applied intranasally during the sensitization and exposure period. Mice were injected intraperitoneally at a dose of 50 mg/kg twice weekly with a total of 12 doses of mAb1 for a period of 6 weeks from 7 weeks to 12 weeks (IL-4 ^hu/hu IL-4Rα ^hu/hu mice , n=12);
(5) HDM sensitization, treated with mouse IL-13Rα2-mFc fusion protein : HDM was applied intranasally during the sensitization and exposure period. Mice were injected intraperitoneally at a dose of 25 mg/kg twice weekly with a total of 12 doses of IL-13Rα2-mFc (IL-4 ^hu/hu IL-4Rα ^hu/hu mice, n=7; wild-type littermates, n=5
);
(6) HDM sensitization, treatment with isotype control antibody : HDM was applied intranasally during the sensitization and exposure period. Mice were injected intraperitoneally at a dose of 50 mg/kg twice a week with a total of 12 doses of isotype control Ab (IL-4 ^hu/hu IL-4Rα ^{hu/ hu} mice, n=7).

Mice were euthanized on day 85, blood was collected for serum immunoglobulin level assays, and lungs (one lobe) were analyzed for i) bronchoalveolar (BAL) lavages, ii) flow cytometry analysis. iii) fixed formalin preparations for staining and histological analysis; or iv) Sir to quantify collagen content per lung lobe.
Used to generate any of the samples for analysis using the col ^™ Collagen Assay.

BAL lavage fluid was obtained from euthanized animals by first exposing the trachea and introducing a 23G lavage tube through a small incision in the tracheal wall. Sterile PBS (1 mL) is then infused into the lungs and the BAL lavage fluid is withdrawn through the lavage tube using a syringe. BAL
100 μL was loaded onto the Cytospin, which was spun at 500 rpm for 5 minutes to extract the cells onto a microscope slide. Slides were dried and H&E stained to visualize eosinophils.

Serum levels of IgE were quantified using a commercial ELISA kit. Briefly, serially diluted serum samples were incubated with anti-IgE capture antibody in 96-well plates and IgE was detected with a biotinylated anti-mouse IgE secondary antibody. HRP-labeled purified mouse IgE was used as a standard.

HDM-specific IgG1 serum levels were quantified by ELISA. Briefly, HDM-coated plates were incubated with serially diluted sera followed by incubation with anti-mouse IgG1 HRP-conjugated antibodies. Relative levels of IgG1 serum levels are expressed as titer units (OD450 multiplied by the dilution factor required to achieve OD450≦0.5). Collected lung lobes were flash frozen in liquid nitrogen and stored at −80° C. until the extraction process. To extract collagen, lungs were homogenized in ice-cold NaCl/NaHCO ₃ solution and centrifuged at 9000×g for 10 minutes. This step was repeated three times and the resulting pellet was digested in acetic acid for 18 hours at 4°C. Samples were centrifuged and supernatants were collected and mixed with Sircol dye reagent to stain for collagen content. Samples were washed with Acid-Salt Wash Reagent to remove unbound Sircol dye and then mixed with Alkali Reagent. 200 μL of each sample was transferred to a 96-well plate and the OD at 555 nm was measured. Collagen standards were used for final quantification of collagen content in each sample.

Lungs were collected from euthanized mice and kept on ice in complete DMEM medium until digested with a mixture of collagenase and DNAse at 37° C. for 20 minutes in HBSS buffer. Collagenase activity was quenched by adding 0.5 M EDTA, samples were centrifuged, and red blood cells were lysed using ACK buffer. The cell suspension obtained for each sample was divided into 3 separate pools and antibody mixture 1 (anti-CD11c-APC Ab, anti-SiglecF-PE Ab, anti-F4/80-FITC Ab, anti-CD45-PerCp-Cy5 .5 Ab), or mixture 2 (anti-CD11c-APC Ab, anti-CD11b-PerCp-Cy5.5 Ab, anti-CD103-FITC Ab, anti-MHCII-PE Ab), or mixture 3 (anti-CD19-PE Ab, anti-Ly6G -APC Ab, anti-CD3-FITC, anti-CD11b-PerCp-Cy5.5 Ab) for 25 minutes at 4°C. Stained cells were fixed with Cytofix/Cytoperm solution for 30 min at 4° C. and stored in PBS until flow cytometry analysis by FACSCanto (BD Biosciences).

From the HDM-induced chronic model of eosinophilic asthma (EA), left lung lobes were harvested from 4 mice per group for microarray analysis of gene expression using GeneChip® technology. Gene expression levels in mice sensitized and exposed with HDM and then treated with isotype control Ab are compared to gene expression levels in mice sensitized and exposed with mock (PBS) and receiving no antibody treatment. bottom. The threshold for changes in gene expression was set to >1.5-fold. The population of genes identified as differentially expressed in HDM-sensitized and exposed mice was then further analyzed in the anti-IL-4Rα treated group compared to the isotype control treated group. The threshold for changes in gene expression in the IL-4Rα-Ab treated group compared to the isotype control treated group was set at >2-fold.

Results HDM sensitization and exposure resulted in increased levels of IgE and HDM-specific IgG1. IgE increases were completely blocked by both anti-IL-4Rα Abs, but not by IL-13Rα2-Fc treatment (FIGS. 27A and 27B); HDM-specific IgG1 levels were not affected by either treatment. did not (data not shown).

HDM sensitization and exposure also caused an increase in collagen content in the lungs of mice. Collagen content in the lungs of mice treated with both IL-4Rα Ab and IL-13Rα2-Fc protein was reduced to levels observed in sham-sensitized and challenged mice (FIGS. 28A and 28B). .

Moreover, mAb1 treatment prevented the influx of eosinophils, neutrophils, and inflammatory dendritic cells into the lung (Figure 29, panels A and B).

Microarray analysis of mRNA isolated from lung tissue of HDM-induced IL-4 ^hu/hu IL-4Rα ^hu/hu mice treated with an isotype control antibody compared to sham-sensitized and sham-exposed mice revealed that 1468 Differential expression of genes (826 up-regulated and 642 down-regulated genes) was revealed. mAb1 treatment of HDM-induced IL-4 ^hu/hu IL-4Rα ^hu/hu mice resulted in expression changes in only 521 genes (sham-sensitized/challenged mice), approximately 65 genes affected by HDM-sensitized/challenged mice. % genes were effectively blocked (>1.5-fold change, p<0.05). mAb1 increased gene expression of several members of the IL-1 cytokine family, specifically IL1α (2.9-fold), IL-33 (2.6-fold) and IL-18 binding protein (1.5-fold) Of particular interest is the finding that it mediated the downregulation of IL-1β gene expression was not increased in the HDM-induced isotype control treated group (compared to sham-sensitized mice), but decreased in the mAb1 treated group (1.5-fold). Gene expression of the Th1 inflammatory cytokines IL-12β and IFN-γ was also downregulated by mAb1 compared to the isotype control treated group. Notably, eight genes encoding chemokine ligands involved in cell homing and trafficking were down-regulated in the mAb1-treated group when compared to the isotype control-treated group: Ccl11 (approximately 9-fold reduction), Ccl8 and Cxcl2 (both Cxcl1, Ccl7, Ccl6 (all about 3-fold decreased), Ccl2 and Ccl9 (about 2-fold decreased).

Conclusions This example demonstrates that blockade of IL-4 signaling through type I and type II receptors by anti-IL-4Rα antibodies was associated with inflammatory and fibrotic changes in the lungs of HDM-exposed mice, as well as HDM-driven genes. It shows that the characteristic change is suppressed.

Other embodiments are within the claims.

Claims (19)

An antibody that specifically binds to the interleukin-4 receptor (IL-4R) or antigen binding thereof for use in reducing the incidence of one or more asthma exacerbations in a subject with bronchial asthma A pharmaceutical composition comprising the fragment,
Said use comprises sequential administration of a single initial dose of said pharmaceutical composition to a subject, wherein administration of said single initial dose is accompanied by one or more secondary doses of said pharmaceutical composition. followed by administration of the dose,
wherein the antibody or antigen-binding fragment thereof comprises three heavy chain complementarity determining region (HCDR) sequences comprising SEQ ID NOs: 148, 150 and 152, respectively, and three light chains comprising SEQ ID NOs: 156, 158 and 160, respectively. comprising a complementarity determining region (LCDR);
A pharmaceutical composition as described above. Asthma exacerbation:
(a) a 30% or greater reduction from baseline in morning peak expiratory flow (PEF) on 2 consecutive days;
(b) 6 or more additional seizure medication puffs of albuterol or levalbuterol (compared to baseline) in 24 hours on 2 consecutive days; and (c) (i) systemic (oral and/or parenteral) steroid treatment, or (ii) an increase in inhaled corticosteroids to at least four times the last dose received prior to discontinuation, or (iii) exacerbation of asthma requiring hospitalization. The pharmaceutical composition of claim 1, wherein An antibody or antigen-binding fragment thereof that specifically binds to the interleukin-4 receptor (IL-4R) for use in improving one or more asthma-related parameters in a subject with bronchial asthma A pharmaceutical composition comprising
Said use comprises sequential administration of a single initial dose of said pharmaceutical composition to a subject, wherein administration of said single initial dose is accompanied by one or more secondary doses of said pharmaceutical composition. followed by administration of the dose,
wherein the antibody or antigen-binding fragment thereof comprises three heavy chain complementarity determining region (HCDR) sequences comprising SEQ ID NOs: 148, 150 and 152, respectively, and three light chains comprising SEQ ID NOs: 156, 158 and 160, respectively. comprising a complementarity determining region (LCDR);
A pharmaceutical composition as described above. Improvements in asthma-related parameters were:
(a) an increase from baseline in forced expiratory volume in 1 second (FEV1) of at least 0.10 L;
(b) an increase from baseline in morning peak expiratory flow (AM PEF) of at least 10.0 L/min;
(c) an increase from baseline in nighttime peak expiratory flow (PM PEF) of at least 1.0 L/min;
(d) reduction from baseline in daily albuterol/lev-albuterol use by at least 1 inhaler/day;
(e) a reduction from baseline of at least 0.5 points in the 5-item Asthma Control Questionnaire (ACQ5) score;
(f) a reduction from baseline of at least 0.2 nightly nocturnal awakenings (number per night) measured daily; 4. The pharmaceutical composition of claim 3, selected from the group consisting of a 5 point reduction from baseline. A pharmaceutical composition comprising an antibody or antigen-binding fragment thereof that specifically binds to the interleukin-4 receptor (IL-4R) for use in increasing FEV1 in litres in a subject with bronchial asthma and
Said use comprises sequential administration of a single initial dose of said pharmaceutical composition to a subject, wherein administration of said single initial dose is accompanied by one or more secondary doses of said pharmaceutical composition. followed by administration of the dose,
wherein the antibody or antigen-binding fragment thereof comprises three heavy chain complementarity determining region (HCDR) sequences comprising SEQ ID NOs: 148, 150 and 152, respectively, and three light chains comprising SEQ ID NOs: 156, 158 and 160, respectively. comprising a complementarity determining region (LCDR);
A pharmaceutical composition as described above. 6. The pharmaceutical composition of any one of claims 1-5, wherein the antibody or antigen-binding fragment thereof is administered to the subject at a dose of about 75 mg to about 600 mg. 6. The pharmaceutical composition of any one of claims 1-5, wherein the antibody or antigen-binding fragment thereof is administered to the subject systemically, intravenously, or intranasally. The pharmaceutical composition of any one of claims 1-5, wherein the antibody or antigen-binding fragment thereof is administered to the subject subcutaneously. 9. The pharmaceutical composition of Claim 8, wherein the antibody or antigen-binding fragment thereof is administered to the subject using an autoinjector, a needle and syringe, or a pre-filled pen delivery device. 6. The pharmaceutical composition of any one of claims 1-5, wherein the antibody or antigen-binding fragment thereof is administered to the subject in combination with a background treatment. 11. The pharmaceutical composition of claim 10, wherein the background treatment is administered to the subject before, after, or at the same time as the antibody or antigen-binding fragment thereof. Background treatments include TNF inhibitors, IL-1 inhibitors, IL-5 inhibitors, IL-8 inhibitors, IgE inhibitors, leukotriene inhibitors, methylxanthines, NSAIDs, nedocromil sodium, cromolyn sodium, 12. The pharmaceutical composition of claim 11, selected from the group consisting of fungal agents, and combinations thereof. 12. The pharmaceutical composition of Claim 11, wherein the background treatment is a systemic steroid. The pharmaceutical composition according to any one of claims 1-5, wherein at least 8 secondary doses are administered. 6. The pharmaceutical composition of any one of claims 1-5, wherein the antibody or antigen-binding fragment thereof comprises the heavy chain variable region (HCVR)/light chain variable region (LCVR) sequence pair of SEQ ID NO: 162/164. . 6. The pharmaceutical composition of any one of claims 1-5, wherein the subject has refractory asthma and the subject's symptoms are not adequately controlled with existing therapies. The pharmaceutical composition of any one of claims 1-5, wherein the subject has eosinophilic asthma. 18. The pharmaceutical composition of claim 17 , wherein the subject has at least 300 cells/[mu]L blood eosinophils and/or at least 3% sputum eosinophils. 6. The pharmaceutical composition of any one of claims 1-5, wherein the subject has elevated levels of exhaled nitric oxide (FeNO).

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